共查询到20条相似文献,搜索用时 46 毫秒
1.
Epidermal growth factor (EGF) at 10 ng/ml reduces by over 50-fold the extracellular Ca2+ required for multiplication of normal human skin fibroblasts. Therefore, a Ca2+-related process may play a central role in the mechanism by which EGF exerts its effect on cell multiplication. 相似文献
2.
The influence of epidermal growth factor (EGF) on the Ca2+ requirement of normal (NP-2s) and neoplastic human epithelial (PC-3) cells was studied using a clonal growth assay. The interaction of Ca2+ and EGF was investigated by kinetic analysis of dose-response experiments in which the Ca2+ or EGF concentration necessary for half-maximal growth, Km were determined. The normal epithelial cells required 80 times more calcium than did the cancer cells. In the presence of EGF, the Ca2+ requirement of both cell types was virtually identical. EGF did not affect the growth rate of the cancer line. The interaction between Ca2+and EGF was found to be unidirectional since EGF reduced the KmCa2+ 120-fold, whereas Ca2+ had no effect on the EGF dose-response. 相似文献
3.
Béatrice Marquèze-Pouey Sébastien Mailfert Vincent Rouger Jean-Marc Goaillard Didier Marguet 《PloS one》2014,9(9)
Signaling mediated by the epidermal growth factor (EGF) is crucial in tissue development, homeostasis and tumorigenesis. EGF is mitogenic at picomolar concentrations and is known to bind its receptor on high affinity binding sites depending of the oligomerization state of the receptor (monomer or dimer). In spite of these observations, the cellular response induced by EGF has been mainly characterized for nanomolar concentrations of the growth factor, and a clear definition of the cellular response to circulating (picomolar) concentrations is still lacking. We investigated Ca2+ signaling, an early event in EGF responses, in response to picomolar doses in COS-7 cells where the monomer/dimer equilibrium is unaltered by the synthesis of exogenous EGFR. Using the fluo5F Ca2+ indicator, we found that picomolar concentrations of EGF induced in 50% of the cells a robust oscillatory Ca2+ signal quantitatively similar to the Ca2+ signal induced by nanomolar concentrations. However, responses to nanomolar and picomolar concentrations differed in their underlying mechanisms as the picomolar EGF response involved essentially plasma membrane Ca2+ channels that are not activated by internal Ca2+ store depletion, while the nanomolar EGF response involved internal Ca2+ release. Moreover, while the picomolar EGF response was modulated by charybdotoxin-sensitive K+ channels, the nanomolar response was insensitive to the blockade of these ion channels. 相似文献
4.
Lowering of extracellular Ca2+ levels will reversibly arrest the growth of human fibroblasts (WI38). Simian virus40(SV40)-transformed WI38 cells do not exhibit this Ca2+-dependent arrest. One possibility for this difference in Ca2+ requirement is that extracellular or surface membrane-bound Ca2+ may be required for growth factor receptor-mediated endocytosis and this Ca2+ requirement may differ in normal versus transformed cells. In this study we have evaluated the role of Ca2+ in the binding, internalization, and degradation of epidermal growth factor (EGF) in the WI38 and SV40 WI38 cell. The binding of [125I]EGF to the cell surface is not significantly altered by lowering of Ca2+ to 10?5-M levels in either the normal or transformed cell. At this Ca2+ level, growth of the normal cell is inhibited. The subsequent internalization of EGF is reduced nearly threefold in the normal cell but not in the transformed cell following Ca2+ deprivation. Degradation of the EGF-receptor complex is also sensitive to Ca2+. A twofold reduction in the rate of release of acid-soluble 125I occurs in the normal but not the transformed cell under conditions of lowered medium Ca2+. In contrast, 2-chloro-10-3-aminopropyl phenothiazine (CP), an inhibitor of the Ca2+-dependent regulator protein calmodulin, causes an inhibition of [125I]EGF internalization and degradation in both the normal and transformed WI38 cell, and a marked inhibition of [125I]EGF binding to the cell surface receptor of the transformed cell but not the normal cell. 相似文献
5.
Calcium,magnesium, and serum factors in multiplication of normal and transformed human lung fibroblasts 总被引:6,自引:0,他引:6
Wallace L. McKeehan Kerstin A. McKeehan 《In vitro cellular & developmental biology. Plant》1980,16(6):475-485
Summary Serum factors determine the extracellular requirement for both Ca2+ and Mg2+ for multiplication of normal human lung fibroblasts in vitro. Serum factors also affect the extracellular Ca2+ requirement for transformed fibroblasts but to a different extent than for normal cells. Transformed cells exhibit a reduced
requirement for both Ca2+ and Mg2+ for multiplication. The apparent reduction in Ca2+ requirement of transformed cells is dependent on the level of serum factors in the medium. The reduced Mg2+ requirement for transformed cells is more striking than the loss of Ca2+ and independent of the level of serum factors in the medium. A sequential effector relationship among serum factors, Ca2+ and Mg2+, in a proliferative control system for normal cells is proposed. Alteration or bypass of an intracellular Mg2+-requiring process is proposed as a major lesion in the transformed cells. This alteration causes an observed loss of requirements
for both Ca2+ and serum factors for the multiplication of transformed cells.
This work was supported by Grant CA-15305 from the National Cancer Institute, Contract 223-74-1156 from the Bureau of Biologics,
Food and Drug Administration, HEW Biomedical Research Support Grant S07RR05800, and the W. Alton Jones Foundation. 相似文献
6.
Peter W. Marks Benjamin A. Kruskal Frederick R. Maxfield 《Journal of cellular physiology》1988,136(3):519-525
The calcium-sensitive fluorescent indicator fura-2 and a microscope equipped for rapidly changing excitation wavelengths were used to look at the effects of growth factors on cytosolic free calcium ([Ca2+]i,) in NRK-49F cells. In these cells bradykinin induced a rapid increase in [Ca2+]i, which generally decayed to near basal [Ca2+]i within 3 minutes. The initial rise in [Ca2+]i in response to bradykinin was relatively independent of extracellular calcium; however, the decay to basal [Ca2+]i was more rapid in the absence of extracellular calcium. Measurements made on individual cells showed a heterogeneity in the response to bradykinin. Epidermal growth factor (EGF) had no effect on [Ca2+]i in NRK-49F cells when added alone in the presence of extracellular calcium. Simultaneous addition of bradykinin and EGF produced a more prolonged increase in [Ca2+]i than bradykinin alone. The prolongation was dependent on the presence of extracellular calcium and did not occur in its absence. Transient increases in [Ca2+]i occurring after the initial peak were occasionally seen in these cells. Our results indicate that there is rapid interaction between the signaling mechanisms for bradykinin and EGF. When this occurs, one effect is the transport of calcium into the cell from the extracellular environment, causing a more prolonged rise in [Ca2+]i. This effect occurs within 1 minute after combined addition of bradykinin and EGF. 相似文献
7.
Summary Calcium signaling systems in nonexcitable cells involve activation of Ca2+ entry across the plasma membrane and release from intracellular stores as well as activation of Ca2+ pumps and inhibition of passive Ca2+ pathways to ensure exact regulation of free cytosolic Ca2+ concentration ([Ca2+]
i
). A431 cells loaded with fura-2 cells were used as a model system to examine regulation of Ca2+ entry and intracellular release. Epidermal growth factor (EGF) and transforming growth factor alpha (TGF-) both stimulated Ca2+ entry and release while bradykinin appeared only to release Ca2+ from intracellular stores. The possible role of protein kinase C (PKC) in modulating the [Ca2+]
i
response to these agonists was examined by four methods. Low concentrations of TPA (2×10–10
m) had no effect on Ca2+ release due to EGF, TGR- or bradykinin but resulted in a rapid return of [Ca2+]
i
to baseline levels for EGF or TGF-. Addition of the PKC inhibitor staurosporine (1 and 10nm)_completely inhibited the action of TPA on EGF-induced [Ca2+]
i
changes. An inhibitor of diglyceride kinase (R59022) mimicked the action of TPA. Down-regulation of PKC by overnight incubation with 0.1 or 1 m TPA produced the converse effect, namely prolonged Ca2+ entry following stimulation with EGF or TGF-. To show that one effect of TPA was on Ca2+ entry, fura-2 loaded cells were suspended in Mn2+ rather than Ca2+ buffers. Addition of EGF or TGF- resulted in Ca2+ release and Mn2+ entry. TPA but not the inactive phorbol ester, 4--phorbol-12,13-didecanoate, inhibited the Mn2+ influx. Thus, PKC is able to regulate Ca2+ entry due to EGF or TGF- in this cell type. A431 cells treated with higher concentrations of TPA (5×10–8
m) inhibited not only Ca2+ entry but also Ca2+ release due to EGF/TGF- but had no effect on bradykinin-mediated Ca2+ release, suggesting differences in the regulation of the intracellular stores responsive to these two classes of agonists. Furthermore, sequential addition of EGF or TGF- gave a single transient of [Ca2+]
i
, showing a common pool of Ca2+ for these agonists. In contrast, sequential addition of EGF (or TGF-) and bradykinin resulted in two [Ca2+]
i
transients equal in size to those obtained with a single agonist. Ionomycin alone was able to fully deplete intracellular Ca2+ stores, whereas ionomycin following either EGF (or TGF-) or bradykinin gave an elevation of the [Ca2+]
i
signal equal to that of the second agonist. These data indicate that there are separate pools of intracellular Ca2+ for EGF-mediated Ca2+ release which also respond differently to TPA. 相似文献
8.
J.L. Hill K. Hammar P.J.S. Smith D.J. Gross 《Molecular reproduction and development》1999,53(2):244-253
Epidermal growth factor (EGF) has received much attention recently for its positive effects on mammalian oocyte maturation and embryo development and its potential importance in cytoplasmic maturation of oocytes. Calcium (Ca2+) homeostasis in germinal vesicle stage oocytes has also been suggested to play a role in cytoplasmic maturation. This study examined the effects of EGF on Ca2+ mobilization as measured by its efflux from mouse oocytes at three time periods throughout maturation (0–4 hr, 4–8 hr, and 12 hr). Immature cumulus oocyte complexes (COCs) removed from the ovary for less than 4 hr exhibit oscillations in Ca2+ efflux that initiated 5–30 min following EGF stimulation. This response was not observed in COCs matured for 4–8 hr or 12 hr or in unstimulated 0–4 hr COCs. Denuded oocytes and cumulus cells did not show the same response to EGF (8.2 nM and 16.4 nM). Immunohistochemistry for detection of the EGF receptor along with EGF internalization studies showed that receptors are present both on cumulus cells and the oocyte but EGF appears to be internalized mainly by the cumulus cells. These data demonstrate that EGF induces oscillations in Ca2+ efflux in COCs 0–4 hr old and this response is mediated by the cumulus cells. Mol. Reprod. Dev. 53:244–253, 1999. © 1999 Wiley‐Liss, Inc. 相似文献
9.
10.
Human neonatal skin fibroblasts plated sparsely in MCDB 105 traversed a complete cell cycle in the absense of serum or serum-derived proteins. Addition of pure PDGF did not significantly increase entrance into S phase as revealed by 3H-thymidine labeling index or clonal growth on palladium islands. In subphysiologic Ca2+ concentrations or in the presence of a calmodulin inhibitor, W7, proliferation in the absence of growth factors ceased and PDGF became mitogenic. In contrast, confluent fibroblast cultures were stimulated by PDGF in physiologic Ca2+ concentrations. This was also the case with sparse adult skin fibroblast cultures while a fetal strain entered S in the absence of PDGF even in low extracellular Ca2+ concentrations. EGF gave similar results as PDGF in all experiments performed. This proposes a similar role for the two growth factors in the cell cycle. However, a difference in the mechanisms of action of PDGF and EGF is indicated by the fact that PDGF and EGF were additive at optimal concentrations when maximal growth response by a single growth factor was restricted by a subphysiologic extracellular Ca2+ concentration. 相似文献
11.
EGF Stimulates Growth by Enhancing Capacitative Calcium Entry in Corneal Epithelial Cells 总被引:2,自引:0,他引:2
Yang H Sun X Wang Z Ning G Zhang F Kong J Lu L Reinach PS 《The Journal of membrane biology》2003,194(1):47-58
In rabbit corneal epithelial cells (RCEC), we determined whether capacitative calcium entry (CCE) mediates the mitogenic response to epidermal growth factor, EGF. [Ca2+]i was measured with single-cell fluorescence imaging of fura2-loaded RCEC. EGF (5 ng/ml) maximally increased [Ca2+]i 4.4-fold. Following intracellular store (ICS) calcium depletion in calcium-free medium with 10 µM cyclopiazonic acid (CPA) (endoplasmic reticulum calcium ATPase inhibitor), calcium addback elicited plasma membrane Ca2+ influx as a result of activation of plasma membrane store operated channel (SOC) activity. Based on Mn2+ quench measurements of fura2 fluorescence, 5 ng/ml EGF enhanced such influx 2.3-fold, whereas with Rp-cAMPS (protein kinase A inhibitor) plus EGF it increased by 5.3-fold. In contrast, SOC activation was blocked with 100 µM 2-aminoethyldiphenylborate (2-APB, store-operated channel inhibitor). During exposure to either 50 µM UO126 (MEK-1/2 inhibitor) or 10 µM forskolin (adenylate cyclase activator), 5 ng/ml EGF failed to affect [Ca2+]i. RT-PCR detected gene expression of: 1) transient receptor potential (TRP) protein isoforms 1, 3, 4, 6 and 7; 2) IP3R isoforms 1–3. Immunocytochemistry, in conjunction with confocal and immunogold electron microscopy, detected plasma membrane localization of TRP4 expression. Inhibition of CCE with 2-APB and/or CPA, eliminated the 2.5-fold increase in intracellular [3H]-thymidine incorporation induced by EGF. Taken together, CCE in RCEC mediates the mitogenic response to EGF. EGF induces CCE through its stimulation of Erk1/2 activity, whereas PKA stimulation suppresses these effects of EGF. TRP4 may be a component of plasma membrane SOC activity, which is stimulated by ICS calcium depletion. 相似文献
12.
Xesus Casabiell Jose L. Zugaza Celia M. Pombo Atanasio Pandiella Felipe F. Casanueva 《Experimental cell research》1993,205(2)
In EGFR-T17 cells, which express high levels of the epidermal growth factor (EGF) receptor, addition of a saturating dose of EGF (10 nM) leads to an increase in Ins(1,4,5)P3/diacylglycerol and also to cytosolic calcium [Ca2+]i due to both intracellular redistribution and influx from extracellular medium. Pretreatment of cells with cis -unsaturated nonesterified fatty acids such as oleic acid (1 to 100 μM) inhibited EGF-stimulated Ins(1,4,5)P3 generation and Ca2+ release from intracellular stores. Furthermore, such a treatment completely suppress Ca2+ influx in a dose-dependent manner. At doses capable of suppressing such early signals, oleic acid did not alter the process of EGF-mediated internalization of the EGF/EGF-receptor complex, suggesting that [Ca2+]i rise did not mediate receptor internalization. EGF-induced cell proliferation assessed by either thymidine incorporation into DNA, direct cell counting, and microscopic observation was not altered by oleic acid, at doses able to block EGF-mediated early signals. In conclusion, suppression of Ins(1,4,5)P3 generation and [Ca2+]i rises by oleic acid did not alter EGF-receptor internalization nor EGF-induced cell mitosis. Such results suggest that [Ca2+]i rise is not instrumental for EGF-stimulated cell proliferation. 相似文献
13.
Sayaka Yoshiki Rie Matsunaga-Udagawa Kazuhiro Aoki Yuji Kamioka Etsuko Kiyokawa Michiyuki Matsuda 《Molecular biology of the cell》2010,21(6):1088-1096
Situated downstream of Ras is a key signaling molecule, Raf1. Increase in Ca2+ concentration has been shown to modulate the Ras-dependent activation of Raf1; however, the mechanism underlying this effect remains elusive. Here, to characterize the role of Ca2+ in Ras signaling to Raf1, we used a synthetic guanine nucleotide exchange factor (GEF) for Ras, eGRF. In HeLa cells expressing eGRF, Ras was activated by the cAMP analogue 007 as efficiently as by epidermal growth factor (EGF), whereas the activation of Raf1, MEK, and ERK by 007 was about half of that by EGF. Using a biosensor based on fluorescence resonance energy transfer, it was found that activation of Raf1 at the plasma membrane required not only Ras activation but also an increase in Ca2+ concentration or inhibition of calmodulin. Furthermore, the Ca2+-dependent activation of Raf1 was found to be abrogated by knockdown of Shoc2, a scaffold protein that binds both Ras and Raf1. These observations indicated that the Shoc2 scaffold protein modulates Ras-dependent Raf1 activation in a Ca2+- and calmodulin-dependent manner. 相似文献
14.
EGF inhibits muscarinic receptor-mediated calcium signaling in a human salivary cell line 总被引:1,自引:0,他引:1
Zhang BX Yeh CK Hymer TK Lifschitz MD Katz MS 《American journal of physiology. Cell physiology》2000,279(4):C1024-C1033
The effects of epidermal growth factor(EGF) on intracellular calcium ([Ca2+]i)responses to the muscarinic agonist carbachol were studied in a humansalivary cell line (HSY). Carbachol (104 M)-stimulated[Ca2+]i mobilization was inhibited by 40%after 48-h treatment with 5 × 1010 M EGF. EGF alsoreduced carbachol-induced [Ca2+]i inCa2+-free medium and Ca2+ influx followingrepletion of extracellular Ca2+. UnderCa2+-free conditions, thapsigargin, an inhibitor ofCa2+ uptake to internal stores, induced similar[Ca2+]i signals in control and EGF-treatedcells, indicating that internal Ca2+ stores were unaffectedby EGF; however, in cells exposed to thapsigargin, Ca2+influx following Ca2+ repletion was reduced by EGF.Muscarinic receptor density, assessed by binding of the muscarinicreceptor antagonistL-[benzilic-4,4'-3HCN]quinuclidinyl benzilate([3H]QNB), was decreased by 20% after EGF treatment.Inhibition of the carbachol response by EGF was not altered by phorbolester-induced downregulation of protein kinase C (PKC) but was enhancedupon PKC activation by a diacylglycerol analog. Phosphorylation of mitogen-activated protein kinase (MAP kinase) and inhibition of thecarbachol response by EGF were both blocked by the MAP kinase pathwayinhibitor PD-98059. The results suggest that EGF decreases carbachol-induced Ca2+ release from internal stores andalso exerts a direct inhibitory action on Ca2+ influx. Adecline in muscarinic receptor density may contribute to EGF inhibitionof carbachol responsiveness. The inhibitory effect of EGF is mediatedby the MAP kinase pathway and is potentiated by a distinct modulatorycascade involving activation of PKC. EGF may play a physiological rolein regulating muscarinic receptor-stimulated salivary secretion. 相似文献
15.
Abstract. Objectives: The gap junction protein, connexin (Cx), plays an important role in maintaining cellular homeostasis and cell proliferation by allowing communication between adjacent cells. Therefore, this study has examined the effect of epidermal growth factor (EGF) on Cx43 and its relationship to proliferation of mouse embryonic stem cells. Materials and methods: Expressions of Cx43, mitogen‐activated protein kinases (MAPKs) and cell cycle regulatory proteins were assessed by Western blot analysis. Cell proliferation was assayed with [3H]thymidine incorporation. Intercellular communication level was measured by a scrape loading/dye transfer method. Results: The results showed that EGF increased the level of Cx43 phosphorylation in a time‐ (≥5 min) and dose‐ (≥10 ng/mL) dependent manner. Indeed, EGF‐induced increase in phospho‐Cx43 level was significantly blocked by either AG 1478 or herbimycin A (tyrosine kinase inhibitors). EGF increased Ca2+ influx and protein kinase C (PKC) translocation from the cytosolic compartment to the membrane compartment. Moreover, pre‐treatment with BAPTA‐AM (an intracellular Ca2+ chelator), EGTA (an extracellular Ca2+ chelator), bisindolylmaleimide I or staurosporine (PKC inhibitors) inhibited the EGF‐induced phosphorylation of Cx43. EGF induced phosphorylation of p38 and p44/42 MAPKs, and this was blocked by SB 203580 (a p38 MAPK inhibitor) and PD 98059 (a p44/42 MAPK inhibitor), respectively. EGF or 18α‐glycyrrhetinic acid (GA; a gap junction inhibitor) increased expression levels of the protooncogenes (c‐fos, c‐jun and c‐myc), cell cycle regulatory proteins [cyclin D1, cyclin E, cyclin‐dependent kinase 2 (CDK2), CDK4 and p‐Rb], [3H]thymidine incorporation and cell number, but decreased expression levels of the p21WAF1/Cip1 and p27Kip1, CDK inhibitory proteins. Transfection of Cx43 siRNA also increased the level of [3H]thymidine incorporation and cell number. EGF, 18α‐GA or transfection of Cx43 siRNA increased 2‐DG uptake and GLUT‐1 protein expression. Conclusions: EGF‐induced phosphorylation of Cx43, which was mediated by the Ca2+/PKC, p44/42 and p38 MAPKs pathways, partially contributed to regulation of mouse embryonic stem cell proliferation. 相似文献
16.
Previous studies have demonstrated a strict extracellular Ca2+ dependence for the G0 to G1 and G1 to S transition in growth factor-treated T51B rat liver cells that is associated with increased levels of protein kinase C activity. Consequently, we have examined these cells for changes in phospholipid-derived second messengers in response to epidermal growth factor (EGF) and thrombin in order to determine which signals are generated during the initiation of the G0 to G1 transition. Thrombin is coupled to a phosphoinositide hydrolyzing phospholipase C, as we have found a rapid Ca2+-independent increase in the levels of inositol 1,4,5-trisphosphate (Ins[1,4,5]P3), inositol 1,4-bisphosphate (Ins[1,4]P2), and inositol 4-monophosphate (Ins[4]P), as well as a concomitant, transient elevation in diacylglycerol. No changes in either intracellular or extracellular choline metabolites, or an increase in DNA synthesis, were found in response to thrombin. By contrast, treatment of T51B cells with EGF results in a slower, more prolonged extracellular Ca2+-dependent increase in both [3H]-glycerol radiolabeled diacylglycerol, and diacylglycerol mass, an increase in choline release into the extracellular medium, and eventually a substantial DNA synthesis. We were, however, unable to detect any changes in phosphatidylinositol (Ptdlns) turnover, either by accumulation of inositol phosphates or by changes in phospholipids in response to EGF. These results indicate that DNA synthesis can readily occur in the absence of stimulated Ptdlns turnover, and that Ptdlns turnover is not sufficient in itself or necessary to induce DNA synthesis and is not necessary for a Ca2+-dependent increase in diacylglycerol. Moreover, we have demonstrated that the extracellular Ca2+-dependent increase in diacylglycerol levels in response to EGF is associated with an increase in extracellular choline release, which is indicative of an activation of a phosphatidylcholine-linked phospholipase D. These results suggest that diacylglycerol sources other than Ptdlns's may be important in the extracellular Ca2+-dependent regulation of EGF-mediated cell replication. © 1995 Wiley-Liss, Inc. 相似文献
17.
The calcium-sensitive forms of adenylyl cyclases (AC) have been revealed in the majority of vertebrate and invertebrate animals,
as well as in several representatives of unicellular organisms, including infusoria. We have found for the first time that
the AC activity in the infusorian Tetrahymena pyriformis changes in the presence of calcium ions. Calcium ions at concentrations of 0.2–20 μM stimulated the activity of this enzyme,
with the maximum of the stimulatory effect being observed at 2 μM Ca2+. At a concentration of 100 μM and higher, the calcium cations inhibited the AC activity. Antagonists of calmodulin W-5 and
W-7 at concentrations of 20–100 μM decreased the stimulatory effect of 5 μM Ca2+, while at the higher concentrations inhibited it completely. Another calmodulin antagonist, chloropromazine, decreased the
Ca2+-stimulated AC activity only at concentrations of 200–1000 μM. The stimulatory effect of serotonin, EGF, and cAMP on AC activity
was enhanced in the presence of 5 μM Ca2+. The stimulatory effect of EGF, cAMP, and insulin on AC was decreased in the presence of 100 μM Ca2+, while the effect of cAMP was also observed in the presence of calmodulin antagonists (500 μM). At the same time, stimulatory
effect of D-glucose did not change in the presence of Ca2+ and calmodulin antagonists. The obtained data indicate that, in the infusorian T. pyriformis, there are calcium-sensitive forms of AC that can be stimulated by EGF, cAMP, insulin, and serotonin. 相似文献
18.
The principles of enzyme kinetic analysis were applied to quantitate the relationships among serum-derived growth factors, nutrients, and the rate of survival and multiplication of human fibroblasts in culture. The survival or multiplication rate of a population of cells plotted against an increasing concentration of a growth factor or nutrient in the medium exhibited a hyperbolic pattern that is characteristic of a dissociable, saturable interaction between cells and the ligands. Parameters equivalent to the Km and Vmax of enzyme kinetics were assigned to nutrients and growth factors. When all nutrient concentrations were optimized and in steady state, serum factors accelerated the rate of multiplication of a normal cell population. The same set of nutrients that supported a maximal rate of multiplication in the presence of serum factors supported the maintenance of non-proliferating cells in the absence of serum factors. Therefore, under this condition, serum factors are required for cell division and play a purely regulatory iole in multiplication of the cell population. The quantitative requirement for 18 nutrients of 29 that were examined was significantly higher (P < 0.001) for cell multiplication in the presence of serum factors than for cell maintenance in the absence of serum factors. This indicated specific nutrients that may be quantitatively important in cell division processes as well as in cell maintenance. The quantitative requirement for Ca2+, Mg2+, K+, Pi, and 2-oxocarboxylic acid for cell multiplication was modified by serum factors and other purified growth factors. The requirement for over 30 other nutrients could not clearly be related to the level of serum factors in the medium. Serum factors also determined the Ca2+, K+, and 2-oxocarboxylic acid requirement for maintenance of non-proliferating cells. Therefore, when either Ca2+, K+, or 2-oxocarboxylic acid concentration was limiting, factors in serum played a role as cell “survival or maintenance” factors in addition to their role in cell division as “growth regulatory” factors. However, with equivalent levels of serum factors in the medium, the requirement for Ca2+, K+, and 2-oxocarboxylic acids was still much higher for multiplication than for maintenance. Kinetic analysis revealed that the concentrations of individual nutrients modify the quantitative requirement for others for cell multiplication in a specific pattern. Thus, specific quantitative relationships among different nutrients in the medium are important in the control of the multiplication rate of the cell population. When all nutrient concentrations were optimal for multiplication of normal cells, the multiplication response of SV40-virus-transformed cells to serum factors was similar to that of normal cells. When serum factors were held constant, transformed cells required significantly less (P < 0.001) of 12 of the 26 nutrients examined. Therefore, the transformed cells only have a growth advantage when the external concentration of specific nutrients limits the multiplication rate of normal cells. Taken together, the results suggest that the control of cell multiplication is intimately related to external concentrations of nutrients. Specific growth regulatory factors may stimulate cell proliferation by modification of the response of normal cells to nutrients. Transforming agents may confer a selective growth advantage on cells by a constitutive alteration of their response to extracellular nutrients. 相似文献
19.
Simon W. Rabkin 《Experimental cell research》1990,188(2)
The purpose of this study was to examine the effect of epidermal growth factor (EGF) on cardiac function and to explore ionic mechanisms as potential explanations for EGF-induced changes in cardiac contractile frequency. Cardiac cell aggregates were prepared from 7-day-old chick embryo hearts and were maintained in culture. EGF over a concentration range of 5 to 20 ng/ ml produced a dose-dependent increase in cardiac contractile frequency. Inhibition of Na+---H+ exchange by amiloride antagonized the action of EGF. Inhibition of Na+---Ca2+ exchange by dichlorobenzamil prevented the effects of EGF. Inhibition of voltage-dependent calcium influx by diltiazem also antagonized the effect of EGF. The positive chronotropic action of EGF was significantly enhanced when the concentration of Na+ or Ca2+ was increased in the medium. These data indicate that EGF has a definite dose-dependent effect on the cardiac contractile frequency that is operative through ionic transport mechanisms that include increased calcium entry through voltage-dependent calcium channels and stimulation of Na+---H+ and Na+---Ca2+ exchange. The similarity in the effects of inhibition of these three ionic mechanisms suggests they are interrelated so that interference at any step in the process inhibits the action of EGF on cardiac myocytes. 相似文献
20.
Junnan Xu Dan Song Qiufang Bai Liping Cai Leif Hertz Liang Peng 《Neurochemical research》2014,39(4):661-667
Glycogenolysis, in brain parenchyma an astrocyte-specific process, has changed from being envisaged as an emergency procedure to playing central roles during brain response to whisker stimulation, memory formation, astrocytic K+ uptake and stimulated release of ATP. It is activated by several transmitters and by even very small increases in extracellular K+ concentration, and to be critically dependent upon an increase in free cytosolic Ca2+ concentration ([Ca2+]i), whereas cAMP plays only a facilitatory role together with increased [Ca2+]i. Detailed knowledge about the signaling pathways eliciting glycogenolysis is therefore of interest and was investigated in the present study in well differentiated cultures of mouse astrocytes. The β-adrenergic agonist isoproterenol stimulated glycogenolysis by a β1-adrenergic effect, which initiated a pathway in which cAMP/protein kinase A activated a Gi/Gs shift, leading to Ca2+-activated glycogenolysis. Inhibition of this pathway downstream of cAMP but upstream of the Gi/Gs shift abolished the glycogenolysis. However, inhibitors operating downstream of the Ca2+-sensitive step, but preventing transactivation-mediated epidermal growth factor (EGF) receptor stimulation, a later step in the activated pathway, also caused inhibition of glycogenolysis. For this reason the effect of EGF was investigated and it was found to be glycogenolytic. Large increases in extracellular K+ activated glycogenolysis by a nifedipine-inhibited L-channel opening allowing influx of Ca2+, known to be glycogenolysis-dependent. Small increases (addition of 5 mM KCl) caused a smaller effect by a similarly glycogenolysis-reliant opening of an IP3 receptor-dependent ouabain signaling pathway. The same pathway could be activated by GABA (also in brain slices) due to its depolarizing effect in astrocytes. 相似文献