Human embryonic kidney cells (HEK-293 cells): characterization and dose-response relationship for modulated release of nerve growth factor for nerve regeneration

The development of engineered constructs to bridge nerve gaps may hold the key to improved functional outcomes in the repair of injured peripheral nerves. These constructs must be rendered bioactive by providing the growth factors required for successful peripheral nerve regeneration. Previous studies demonstrated that harvested human and rat dermal fibroblasts could be genetically engineered to release nerve growth factor (NGF) both in vitro and in vivo. The use of fibroblasts, however, has the potential to cause scarring, and the expression of NGF from those cells was transient. To overcome these potential difficulties, human embryonic kidney cells were modified for use with the ecdysone-inducible mammalian expression system. These cells (hNGF-EcR-293) have been engineered and regulated to secrete human NGF in response to the ecdysone analogue ponasterone A. HEK-293 cells were transfected with human NGF cDNA with the ecdysone-inducible mammalian expression system (Invitrogen, Carlsbad, Calif.). Stable clones were then selected. Ponasterone A, an analogue of ecdysone, was used as the inducing agent. The secretion of NGF into the medium was analyzed with two different methods. After 24 hours of exposure to the inducing agent, cell medium was transferred to PC-12 cells seeded in 12-well plates, for determination of whether the secreted NGF was bioactive. Medium from untreated or ponasterone A-treated hNGF-EcR-293 cells was deemed bioactive on the basis of its ability to induce PC-12 cell differentiation. The concentrations of secreted NGF were also quantified with an enzyme-linked immunosorbent assay, in triplicate. NGF production was measured in successive samples of the same medium during a 9-day period, with maximal release of 9.05 +/- 2.6 ng/ml at day 9. Maximal NGF production was 8.46 +/- 2.1 pg/10(3) cells at day 9. These levels were statistically significantly different from levels in noninduced samples (p 相似文献   

PC-12 cells hydrolyze the fibrin clot by producing both plasminogen and its tissue activator

It has been shown that rat pheochromocytoma PC-12 cells degraded fibrin clots in vitro. SDS-PAGE performed in the gels of different density and Western blotting using monoclonal antibodies against DD- and D-fibrin fragments demonstrated that both high and low molecular weight degradation products similar to those of fibrin hydrolysis by plasmin had been formed. Enzyme electrophoresis, chromogenic assay and enzyme-linked immunosorbent assay using tPA-specific antibodies demonstrated that PC-12 cells constitutively secreted both plasminogen and tPA. The results obtained allow using PC-12 cells as a model to examine the interaction of nerve cells with the fibrin clot.     

Evidence for a lack of functional receptors for nerve growth factor (NGF) in chick bone cells in vitro

Nerve growth factor (NGF) is essential for the development and differentiation of sympathetic and sensory neurons. Recently, NGF receptors were demonstrated in non-neural cells, and several mesenchymal cell types including lymphocytes and skeletal myotubes were shown to be stimulated to proliferate by NGF. Our purpose was to examine for the presence of functional NGF receptors in osteoblasts. Bone cells from chick calvaria were used as a model; PC-12 cells derived from rat adrenal pheochromocytoma were used as positive controls. NGF was examined for functions in chick bone cells by studying effects on (1) [³H]-thymidine incorporation; (2) alkaline phosphatase (ALP) activity; and (3) protein tyrosine phosphorylation. Effects of NGF on thymidine incorporation and protein tyrosine phosphorylation by PC-12 cells were also measured. A radioreceptor assay was used to test for the presence of receptors. In chick calvarial cells, NGF had no effect on thymidine incorporation, ALP activity or protein tyrosine phosphorylation. Radioreceptor assay with bone cells showed no evidence of NGF receptors. In contrast, in PC-12 cells, NGF (1) decreased thymidine incorporation; (2) increased protein tyrosine phosphorylation; and (3) showed receptor activity by radioreceptor assay. In conclusion, unlike several other mesenchymal cell types, chick bone cells show no evidence of NGF receptors or functional responses to NGF in vitro.     

Quantitative analysis of nerve growth factor in the amniotic fluid during chick embryonic development

Nerve growth factor (NGF) and most neurotrophic factors support the proliferation and survival of particular types of neurons. Besidesthe pivotal role of NGF in the development of neuronal cells, it also has important functions on non-neuronal cells. The amnion surrounds the embryo, providing an aqueous environment for the embryo. A wide range of proteins has been identified in human amniotic fluid (AF). In this study, total protein concentration (TPC) and NGF level in AF samples from chick embryos were measured using a Bio-Rad protein assay, enzyme linked immunosorbent assay (ELISA) and Western blot. TPC increased from days E10 to day E18. There was a rapid increase in AF TPC on day E15 when compared to day E16. No significant changes in NGF levels have been seen from day E10 to day E14. There was a rapid increase in NGF content on days E15 and E16, and thereafter the levels decreased from day E16 to day E18. Since, NGF is important in brain development and changes in AF NGF levels have been seen in some CNS malformations, changes in the TPC and NGF levels in AF during chick embryonic development may be correlated with cerebral cortical development. It is also concluded that NGF is a constant component of the AF during chick embryogenesis.     

Sea urchin hyalin: appearance and function in development

Embryonic chicken sensory cells from dorsal root ganglia and a clonal line of pheochromocytoma cells (PC-12) extended neuronal-like processes within 24 hr of seeding on a naturally produced, basement membrane-like extracellular matrix (ECM) in the absence of nerve growth factor (NGF). Plating on ECM also induced a rapid cell attachment and flattening of these cells and supported the survival of embryonic sensory cells in primary cultures. Unlike the effect of NGF on PC-12 cells, the ECM-induced morphological differentiation was transient and led to disintegration and degeneration of processes bearing PC-12 cells. The ECM-induced morphological differentiation was not inhibited by anti-NGF antibodies, and the cells retained their ability to bind and internalize NGF in a manner similar to that observed on plastic. PC-12 cell attachment and flattening occurred on dishes coated with collagen type IV in a way similar to that observed on ECM, but precoating the dishes with fibronectin had no effect. Extension of cell processes was not induced by either substrate. Morphological differentiation but not the induction of cell adhesion and flattening was inhibited by either prefixation with glutaraldehyde, oxidation with periodate, or preexposure to concanavalin A of the ECM, suggesting that the ECM and in particular its sugar moieties play an active role in the induction of neurite outgrowth. It is suggested that close contact with the ECM provides chemical or mechanical cues that permit contactmediated elongation and directed growth of both embryonic and regenerating nerve fibers.     

Transcriptional regulation of early growth response genes in FOS-expressing PC-12 cells.

Deregulated c-fos expression in the rat pheochromocytoma cell line, PC-12, causes pronounced downregulation of nerve growth factor (NGF)-induced c-fos and c-jun activation, accompanied by a block in NGF-induced differentiation of PC-12 cells. The FOS-expressing PC-12 cells were exposed to diverse agents such as NGF, epidermal growth factor (EGF), basic fibroblast growth factor (bFGF), interleukin-1 beta (IL-1 beta), interleukin-6 (IL-6), dibutyryl cyclic adenosine 3',5' monophosphate (db cAMP), and Ca-ionophore; and the expression of egr-1, c-fos, c-jun, jun-B, and jun-D was analyzed. Pronounced downregulation of c-fos, c-jun, and--to a lesser extent--jun-B was observed on treatment with NGF, bFGF, db cAMP, and Ca-ionophore, whereas EGF-induced activation of these early response genes was not inhibited in FOS-expressing PC-12 cells. Ca-ionophore- and db cAMP-induced egr-1 activation in PC-12 fos cells was completely inhibited. Both parent and PC-12 fos cells expressed similar high basal levels of jun-D, whose expression was the least regulatable by all of these agents. Transfection of fos promoter-chloramphenicol acetyltransferase (promoter-CAT) plasmid into these stable FOS-expressing PC-12 cells revealed that these effects are exerted at the fos promoter level.     

Qualitative and quantitative analyses of Epstein-Barr virus early antigen diffuse component by western blotting enzyme-linked immunosorbent assay with a monoclonal antibody.

We report the use of monoclonal antibody against the early antigen diffuse component (anti-EA-D) of Epstein-Barr virus (EBV) to analyze, both qualitatively and quantitatively, the expression of EA-D in various human lymphoblastoid cell lines activated by chemical inducers. The kinetics of synthesis of EA-D in P3HR-1, B95-8, and Ramos/AW cells were similar in that they all reached the peak of synthesis on day 5 after induction. Surprisingly, no expression of EA-D was found in induced BJAB/GC, an EBV-genome-containing cell line. EBV-negative cell lines, BJAB and Ramos, were negative for EA-D. Raji cells had no detectable EA-D but responded rapidly to induction, reaching a peak on day 3. Superinfection of Raji cells also resulted in marked induction of EA-D, which reached a plateau between 8 to 12 h postinfection. Western blotting coupled with the enzyme-linked immunosorbent assay was employed to identify polypeptides representing EA-D. A family of four polypeptides with molecular weights of 46,000 (46K protein), 49,000, 52,000, and 55,000 were identified to be reactive with monoclonal anti-EA-D antiserum. The pattern of EA-D polypeptides expressed in each cell line was different. Of particular interest was the expression of a large quantity of 46K protein both in induced Raji and P3HR-1 cells, but not in superinfected Raji cells. A 49K doublet was expressed in activated p3HR-1, B95-8, and Ramos/AW cells and in superinfected Raji cells. In addition, two distinct 52K and 55K polypeptides were expressed in induced Ramos/AW and superinfected Raji cells. However, none of these EA-D polypeptides was detectable in BJAB/GC, BJAB, Ramos, and mock-infected Raji cells. To approximate relative concentrations of EA-D in cell extracts, we employed the enzyme-linked immunosorbent assay and immunoblot dot methods by using one of the purified EA-D components to construct a standard curve. Depending upon the cell lines, it was estimated that ca. 1 to 3% (determined by the enzyme-linked immunosorbent assay) and 0.8 to 1.6% (determined by immunoblot dot) of total proteins from maximally induced cells were EA-D. These results suggest that differential expression of EA-D polypeptides could be of importance in the diagnosis of state of EBV infection.     

trans activation of nerve growth factor in transgenic mice containing the human T-cell lymphotropic virus type I tax gene.

Three lines of transgenic mice containing the human T-cell lymphotropic virus type I (HTLV-I) tax gene develop neurofibromas composed of perineural fibroblasts (S. H. Hinrichs, M. Nerenberg, R. K. Reynolds, G. Khoury, and G. Jay, Science 237:1340-1343, 1987; M. Nerenberg, S. H. Hinrichs, R. K. Reynolds, G. Khoury, and G. Jay, Science 237:1324-1327, 1987). Tumors and tumor cell lines derived from these mice produce neurite outgrowth from PC-12 cells and nerve growth factor (NGF), as determined by RNA (Northern) blot analysis and enzyme-linked immunosorbent assays. In vitro cotransfection studies demonstrate that Tax is able to trans activate the NGF promoter in NIH 3T3 fibroblast cells. The major cis-acting tax-responsive element in the NGF promoter (AGGGTGTGACGA) has 92% homology with a tax-responsive element contained within the 21-bp repeats of the HTLV-I long terminal repeat. The receptor for NGF is also expressed in the transgenic tumor cells, suggesting that Tax may activate an autocrine mechanism through the upregulation of NGF.     

Lack of correlation between 3-hydroxy-3-methylglutaryl coenzyme A reductase activity and lovastatin resistance in nerve growth factor treated PC-12 cells

Summary 1. The relationships among the mevalonic acid (MVA) forming enzyme, 3-hydroxy-3-methylglutaryl coenzyme A (CoA) reductase, cell growth and differentiation, and the cytotoxic effects of the reductase inhibitor lovastatin were studied in PC-12 cells, exposed to growth factors.2. When added individually, nerve growth factor (NGF), basic fibroblast growth factor, and epidermal growth factor induce an increase in HMG-CoA reductase activity in cells grown in serum-containing medium. In the presence of serum, the effect of NGF on HMG-CoA reductase is persistent.3. Short-term serum starvation and long-term NGF treatment, in combination, have an additive effect, resulting in a high reductase activity.4. Unlike serum and MVA, which downregulate levels of HMG-CoA reductase by accelerating its degradation, NGF upregulates reductase by slowing the rate of its degradation. This mechanism, however, appears to operate only in the presence of serum, as after prolonged growth with NGF in serum-free medium, cells have a low reductase activity.5. PC-12 cells grown in the absence of NGF are highly sensitive to lovastatin (25 µM) and more than 70% of the cells die after 48 hr. NGF confers lovastatin resistance on cells grown in the presence or in the absence of serum (only 30–40% cell death after 48 hr with lovastatin).6. NGF-induced resistance on lovastatin develops with time and is apparent only in the well-differentiated PC-12 cells whether or not the cells express a high reductase activity.7. Thus, levels of HMG-CoA reductase activity and lovastatin resistance in PC-12 cells are not directly correlated, though clearly inversed lovastatin cytotoxicity and elevated reductase activities are expressed during the period of cell proliferation.8. These data suggest that fully differentiated neuronal cells may not be affected by prolonged high doses of lovastatin.     

Probable Mechanisms of the Effects of Cerebral on Morphological Characteristics of Rat Pheochromocytoma Cells

Effects of a recently proposed organic preparation, Cerebral, used in the treatment of some vascular brain pathologies, were examined on cultured rat pheochromocytoma PC-12 cells. The normal culture media in six of the seven groups of cell cultures were replaced by similar media with the addition of 0.2 or 2.0 mg/ml Cerebral, 400 mg/ml nerve growth factor (NGF), 1.0 μM verapamil, a blocker of high-threshold calcium channels, or combinations of the above Cerebral concentrations with 1.0 μM verapamil. Within six days of the test period, we measured mean values of the density of cultured cells, proportions of the units possessing significant processes, and projective areas of the cell somata; according to the latter parameter, conventional diameters of the cells (diameters of the circles equivalent to the above area) were calculated. Culturing of PC-12 cells under control conditions (in the non-modified medium) was not accompanied by transformation of their overwhelming majority into neuron-like units. Cerebral in both tested concentrations significantly suppressed proliferation of PC-12 cells, intensified the formation of processes (some of which demonstrated a typical neurite-like structure), and led to an increase in the dimension of the cells. Under conditions of the action of NGF, similar effects were observed, but their intensity was greater. Manifestations of cell differentiation induced by Cerebral developed with a greater delay and were smoothed earlier, as compared with the respective NGF effects. The addition of 1.0 μM verapamil to the culture medium promoted the process of cell differentation; the effects observed were comparable with results of the action of Cerebral. Combinations of verapamil with Cerebral in both above-mentioned concentrations provided somewhat more intense modulatory effects on PC-12 cells than isolated applications of Cerebral, but the effects observed were not additive. The probable mechanisms of changes in the morphological characteristics of PC-12 cells under the influence of Cerebral and the dependence of such changes on the state of calcium signalization are discussed. The data obtained agree with a supposition that the active substances of Cerebral correspond to trophinotropins initiating and/or intensifying synthesis of NGF.     

IgG anti-tetanus toxoid antibody production induced by Epstein-Barr virus from B cells of human marrow transplant recipients

This investigation shows that Epstein-Barr virus (EBV)-activated human B cells from marrow transplant recipients can produce in vitro IgG anti-tetanus toxoid antibody (anti-TT) without booster immunizations with tetanus toxoid (TT). Purified B cells (E-rosette negative) from 8 normal subjects, 6 healthy long-term marrow graft recipients, and 15 long-term marrow graft recipients with chronic graft-vs-host disease (GVHD), were stimulated for 12 days with EBV to induce anti-TT production in culture supernatants. The amount of anti-TT in culture supernatants was quantitated using a enzyme-linked immunosorbent assay. B cells from all 8 normal controls produced in vitro IgG anti-TT after EBV stimulation. Five of 6 healthy recipients had B cells that produced anti-TT after EBV stimulation. Four of 15 recipients with chronic GVHD had B cells capable of producing anti-TT after EBV stimulation. The number of cultures making anti-TT responses was less in those with chronic GVHD than in those without chronic GVHD or normal individuals (P less than 0.001). B cells from patients with chronic GVHD had fewer responses exceeding the overall median of 0.7 ng/ml when compared with the other two groups (P less than 0.03). These data show that B cells of donor origin can produce in vitro IgG anti-TT antibody to tetanus toxoid antigen in a T-independent fashion.     

Nerve growth factor stimulates the activities of the raf-1 and the mitogen-activated protein kinases via the trk protooncogene.

Nerve growth factor (NGF) binds to two structurally unrelated transmembrane proteins on the surface of PC-12 cells, a 75-kDa glycoprotein with a short cytoplasmic sequence, and the trk protooncogene (pp140c-trk), a protein tyrosine kinase activated by NGF. Immediately after binding to cells, NGF induces changes in serine/threonine phosphorylation of several proteins. We have explored the relative roles of these two NGF binding proteins in mediating the activation of two intracellular kinases that may be responsible for some of these phosphorylations. The raf-1 protooncogene is a serine/threonine kinase activated by several growth factors and oncogenic proteins. Treatment of PC-12 cells with NGF increases the serine and threonine phosphorylation of raf-1 in an anti-raf-1 immunoprecipitate kinase assay. This increased phosphorylation observed in vitro is dose-dependent and transient and is accompanied by the NGF-dependent shift in the mobility of immunoblotted raf-1 on SDS sodium dodecyl sulfate-polyacrylamide gel electrophoresis, an effect thought to reflect phosphorylation. NGF-dependent activation of raf-1 is not dependent on protein kinase C, since prolonged exposure to phorbol esters under conditions that cause down-regulation of cellular protein kinase C activity has no effect on the NGF response. Expression of pp140c-trk in 3T3 fibroblasts (3T3-c-trk), as evidenced by cross-linking of 125I-NGF to the 140-kDa protein, permits the NGF-dependent activation of raf-1 kinase, detected in the immunoprecipitate kinase assay, anti-raf immunoblot shift on gel electrophoresis, and incorporation of [32P]orthophosphate into the raf-1 protein. The concentration dependence of raf-1 activation is identical in 3T3-c-trk and PC-12 cells, despite the absence of the 75-kDa NGF binding protein in 3T3-c-trk cells. NGF is without effect in untransfected 3T3 cells or in Chinese hamster ovary cells overexpressing p75, although raf-1 is present in these cells. Similarly, the NGF-dependent activation of mitogen-activated protein (MAP) kinase is detected in 3T3-c-trk cells, but not in untransfected 3T3 or Chinese hamster ovary cells overexpressing p75. As described for raf-1 activation, the NGF dose responses for MAP kinase activation in 3T3-c-trk and PC-12 cells are virtually superimposable. These data indicate that the activation of these two serine/threonine kinases by NGF is mediated solely by binding to and activating the pp140c-trk receptor.     

Mucosal vaccination with a recombinant OprF-I vaccine of Pseudomonas aeruginosa in healthy volunteers: comparison of a systemic vs. a mucosal booster schedule

We compared the immunogenicity of two vaccination schedules with either a systemic or a mucosal booster, both following a mucosal primary vaccination with a recombinant outer membrane fusion protein of Pseudomonas aeruginosa (OprF-I) in 12 healthy volunteers. The systemic booster induced higher levels of OprF-I-specific serum antibodies of IgG isotype, with a mean+/-S.E.M. of 32.6+/-7.8x10(7) enzyme-linked immunosorbent assay (ELISA) units (EU) as compared to the nasal booster with 14.6+/-2.1x10(7) EU (P=0.05). Specific serum IgA antibodies and antibodies in saliva did not differ between the two vaccination groups. We conclude that a combined mucosal/systemic vaccination with the OprF-I vaccine may offer an enhanced systemic immunogenicity. Further studies on the long-term immunogenicity and induction of antibodies on the respiratory airway surface are warranted.     

An Enzyme-Linked Immunoassay for Nerve Growth Factor (NGF): A Tool for Studying Regulatory Mechanisms Involved in NGF Production in Brain and in Peripheral Tissues

A sensitive enzyme-linked immunosorbent assay (ELISA) for nerve growth factor (NGF) has been developed. The sensitivity of this assay (0.1 pg/well) permits the quantification of endogenous immunoreactive NGF in the peripheral nervous system and the CNS. Studies on the regulatory mechanisms involved in NGF production indicate that, in addition to neurally mediated mechanisms, other stimuli, e.g., inflammation, significantly contribute to NGF production.     

Cholera Toxin and Dibutyryl Cyclic AMP Inhibit the Expression of Neurofilament Protein Induced by Nerve Growth Factor in Cultures of Naive and Primed PC12 Cells

The effects of nerve growth factor (NGF), dibutyryl cyclic AMP (db cAMP), and cholera toxin on neurofilament protein expression in cultures of PC12 rat pheochromocytoma cells were examined using an enzyme-linked immunoadsorbent assay (ELISA). Morphological differentiation induced by NGF was associated with up to 30-fold increases in the level of neurofilament protein recognised by monoclonal antibody RT97. A more rapid response was apparent from primed as compared to naive PC12 cells. Cholera toxin and db cAMP both induced morphological differentiation of naive PC12 cells, but failed to promote neurite regeneration from primed cells. Neither response was associated with a significant induction of neurofilament protein. Both cholera toxin and db cAMP, but not B-cholera toxin nor antibodies to the toxin receptor, were found to inhibit the neurofilament protein response induced by NGF. Primed cells were more susceptible to this inhibition, and both cholera toxin and db cAMP inhibited neurite regeneration from these cells. These data suggest that increased intracellular cyclic AMP can suppress the expression of neuronal differentiation antigens induced by NGF, and are consistent with a role for neurofilament protein in promoting or facilitating the formation of a stable neuritic network.     

Tumor promoter modulation of epidermal growth factor- and nerve growth factor-induced adhesion and growth factor binding of PC-12 pheochromocytoma cells

Nerve growth factor (NGF) has previously been shown to increase the rate of adhesion of PC-12 pheochromocytoma cells to cell culture dishes. This increase in the rate of adhesion was postulated to be important in NGF-mediated neurite outgrowth. We now report that epidermal growth factor (EGF) is also able to increase the rate of adhesion of PC-12 cells to cell culture dishes, but does not elicit neurite outgrowth. The dose-response curve for EGF is bell-shaped, in contrast to the more classically shaped dose-response curve obtained with NGF. Tetradecanoyl-phorbol-acetate (TPA), a potent tumor promoter, blocks the EGF-induced increase in adhesion rate of PC-12 cells, but does not alter the NGF-induced increase in adhesion rate. TPA shifts the EGF binding curve to the right for PC-12 cells, but does not alter maximal EGF binding at saturating concentrations of EGF. The binding of NGF to PC-12 cells is not affected by TPA. NGF-induced neurite formation by PC-12 cells is unaffected by TPA, in contrast to the previously reported delay of neurite outgrowth of serum-deprived neuroblastoma cells and NGF-exposed chick embryonic ganglia cells. NGF and EGF both cause a decrease in the number of short microvilli and an increase in the number of long microvilli on PC-12 cells. TPA blocks the decrease in the number of short microvilli in EGF-treated cells, but not in NGF-treated cells. Long microvilli formation is blocked by TPA in both conditions, suggesting the latter are not involved in the increased adhesion rates.     

眼镜蛇毒神经生长因子抑制肝星状细胞增殖并诱导其凋亡

目的:从眼镜蛇毒中分离纯化神经生长因子(Nerve Growth Factor,NGF),观察眼镜NGF对肝星状细胞HSC-T6增殖、凋亡活性的影响,进一步为蛇毒NGF在抗肝纤维化治疗提供依据。方法:采用shephadex G-75和CM Sepharose CL-6B二步柱色谱对眼镜蛇毒NGF进行纯化分离;PC12细胞测定各洗脱峰的活性,再用SDS-PAGE鉴定具有NGF活性洗脱峰的纯度和相对分子质量。实验设立空白对照和NGF处理组,分别作用于HSC-T6,培育相应时间,MTT检测眼镜蛇毒NGF对HSC-T6细胞活力影响;HE染色、紫外激光显微镜与透射电镜观察HSC-T6细胞的形态学变化;TUNEL、流式细胞技术检测眼镜蛇毒NGF对HSC-T6细胞凋亡的影响。结果:眼镜蛇毒经PC-12细胞鉴定第Ⅵ峰具有NGF活性;SDS-PAGE检测为电泳纯,相对分子质量为22.3KD;NGF对HSC-T6细胞增殖具有明显抑制作用(2μg/ml NGF的抑制率为49.66%±6.50%,P<0.05;6.25μg/ml NGF的抑制率为71.33%±1.53%,P<0.05);TUNEL法检测发现NGF干预组的凋亡率28.71%±1.59%(2ug/ml NGF)和34.4%±2.49%(5μg/mlNGF)明显高于对照组的15.85%±1.58%(P<0.05);流式细胞仪也有同样的发现,NGF干预组的凋亡率16.12%±3.02%(2 ug/mlNGF)和21.15%±3.31%(5μg/ml NGF)明显高于对照组的2.7%±1.55%(P<0.05)。结论:眼镜蛇毒NGF能抑制肝星状细胞HSC-T6增殖并诱导其凋亡。     

The Effects of Rosuvastatin on the Serum Cortisol,Serum Lipid,and Serum Mevalonic Acid Levels in the Healthy Indian Male Population

In this open-label, balanced, randomized, placebo-controlled, parallel study, healthy male volunteers were randomly divided into two groups. Each group received either a single oral dose of rosuvastatin 20 mg or placebo. Estimations were done at predose on day 1 of dosing (baseline) and 24 h postdose after days 7 and 14. Serum cortisol and serum lipid levels were estimated using enzyme-linked immunosorbent assay kits and serum mevalonic acid (MVA) levels were measured using validated liquid chromatography–tandem mass spectrometry method. Rosuvastatin produced a statistically significant (P < 0.05) decrease in total cholesterol, low-density lipoprotein cholesterol, very low-density lipoprotein cholesterol, and triglycerides. However, the increase in high-density lipoprotein cholesterol and decrease in cortisol and MVA were not statistically significant when compared to the placebo-treated group. The study showed that rosuvastatin at a dose of 20 mg/day for a period of 14 days was very potent as cholesterol-lowering agent, without any significant change in serum cortisol level in the healthy Indian male population.     

TRPV2激活对缺氧再灌注时体外培养星形胶质细胞神经生长因子释放的影响

神经生长因子对脑缺血后神经元的存活有重要意义。该研究观察了TRPV2激活剂2APB对体外缺血再灌注模型中原代培养大鼠大脑皮层星形胶质细胞神经生长因子释放的影响。将原代培养大鼠大脑皮层星形胶质细胞分为2APB组（0．5mmol／L）和对照组（不含2APB）,在糖氧剥夺情况下培养2h,然后恢复正常全培养基复氧培养48h。用Westem blot检测星形胶质细胞神经生长因子的表达水平;用ELISA检测星形胶质细胞条件培养液中神经生长因子的含量。结果表明,0．5mmol／L2APB可以诱导正常情况下及糖氧剥夺再灌注情况下体外培养星形胶质细胞NGF的合成和释放LP〈0．01）。此外,JNK阻滞剂可抑制糖氧剥夺再灌注情况下2APB诱导的星形胶质细胞神经生长因子的释放。综上．TRPV2激活可以影响糖氧剥夺再灌注情况下体外培养星形胶质细胞神经生长因子的合成和释放。TRPV2有可能成为脑缺血再灌注后的潜在治疗靶点。