Impaired synergism of dexamethasone and IGF-I in cultures of aged fibroblasts.
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Aging↗
Biomedical subjects
P D Phillips
Publications and source records attributed to P D Phillips.
Impaired synergism of dexamethasone and IGF-I in cultures of aged fibroblasts.
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Cell Survival↗
Insulin-like growth factor-I: specific binding to high and low affinity sites and mitogenic action throughout the life span of WI-38 cells.
Insulin-like growth factor-I (IGF-I) (13 nM) can replace insulin (0.8 microM) in a serum-free medium containing epidermal growth factor (EGF) (16 nM) and dexamethasone (DEX) (140 nM) and stimulate DNA synthesis in young cultures of WI-38 cells, similar to the stimulation of serum-supplemented medium. By contrast, senescent cells become unresponsive to all of these hormones. The effect of IGF-I, EGF, and DEX is synergistic in stimulating multiple rounds of low density cell division. Total specific binding of [125]IGF-I per cell in monolayer culture does not change with age, which indicates, in light of increased cell size with age, an actual decrease in specific binding per micron2 of cell surface area. Binding can be traced to two separate cell proteins. Binding to the alpha subunit of the IGF-I transmembrane receptor may increase slightly with age while the 50% displacement remains unchanged. The remainder of the IGF-I specific binding (five- to thirty-fold more) is to a low molecular weight, cell-associated binding protein whose 50% displacement is 10 times higher, but also remains unchanged with age. Specific binding to the lower affinity sites decreases slightly with age at equal cell densities. IGF-I binding to the alpha subunit of the transmembrane receptor is independent of cell density, while binding to the low molecular weight binding protein is inversely proportional to cell density and may vary by as much as tenfold.
Cell Division↗
EGF-dependent phosphorylation of the EGF receptor in plasma membranes isolated from young and senescent WI-38 cells.
Tyrosine-specific phosphorylation of the receptor for epidermal growth factor (EGF) in plasma membranes isolated from WI-38 cells is EGF-dependent and occurs to an equivalent extent and on identical tryptic peptides in preparations from cells of various in vitro ages. There is a marked reduction, however, in phosphorylation of receptor molecules from senescent as compared with young WI-38 cells, if enzyme activity is assayed in an immune complex following solubilization of plasma membranes with Nonidet P-40 (NP-40). Differences in the level of receptor phosphorylation in young vs. senescent NP-40 extracts are not resolved by changing the temperature at which the assay is performed, or the length of incubation. Moreover, addition of NP-40 or chloroform-methanol extracts of young cells to assays measuring receptor phosphorylation in senescent cell NP-40 preparations does not augment the senescent enzyme activity. The immunopurified senescent receptor is, however, capable of catalyzing phosphorylation of exogenous substrates. These results indicate that the loss of receptor autophosphorylation in solubilized preparations may result from a differential sensitivity of the senescent cell receptor to the detergent. This finding provides a marker for senescence and suggests subtle changes in protein structure, conformation, or regulation of the EGF receptor in senescent cells.
Cell Line↗
Day-case herniotomy in children. A comparison of ilio-inguinal nerve block and wound infiltration for postoperative analgesia.
Forty-nine boys scheduled for day-case inguinal herniotomy were studied to compare ilio-inguinal nerve block and wound infiltration for postoperative analgesia. Both techniques were simple to perform and produced no complications. In the ilio-inguinal block group, 100% had either no pain or very mild discomfort when assessed 60 minutes after return to the day unit, compared to 95% in the infiltration group. Some children did appear to have pain following discharge but in all cases this responded well to simple analgesics. We conclude that both techniques provide satisfactory analgesia whilst the complications of narcotics are avoided, and suggest that simple infiltration of the wound with local anaesthetic solution should be encouraged in paediatric anaesthesia.
Ambulatory Surgical Procedures↗
Phenotypic variability among density-fractionated human lung fibroblasts.
To determine if there is phenotypic variability within apparently homogeneous human lung fibroblast lines, growth-arrested lung fibroblasts were separated according to cell density. Three fibroblast fractions of specific gravity 1.023 to 1.035, 1.036 to 1.049, and 1.052 to 1.059 were obtained. These fractions differed in their cell volume and proliferative response to serum, but not in their response to growth-inhibiting soluble factors from lung mononuclear cells. The differences in proliferative capacity were not maintained during in vitro culture, and were not the result of the fractionation procedure selecting for cells at a particular point in the cell cycle. However, the fractionation procedure did separate cells based in part on their propensity to traverse the cell cycle. The least dense fraction was enriched for larger cells that were noncycling or slowly cycling, whereas the densest fibroblast fraction was enriched for small, rapidly cycling cells. Thus, there is morphologic and functional variability within adult human lung fibroblast lines. This phenotypic variability may be an important factor controlling the pulmonary fibrotic response.
Cell Cycle↗
A dynamic traction splint for the management of extrinsic tendon tightness.
The dynamic traction splint designed by therapists at Walter Reed Army Medical Center is used for the management of extrinsic extensor tendon tightness commonly seen in brachial plexus injuries and traumatic soft tissue injuries of the upper extremity. The two components of the splint allow for simultaneous maximum flexion of the MCP and IP joints. This simple and economical splint provides an additional modality to any occupational therapy service involved in the management of upper extremity disorders.
Finger Joint↗
The effect of dexamethasone on epidermal growth factor binding and stimulation of proliferation in young and senescent WI38 cells.
Addition of 0.14 microM dexamethasone (DEX) to young log-phase WI38 cultures seeded at various densities in serum-free medium containing 4.1 nM epidermal growth factor (EGF) resulted in a synergistic increase in proliferation and final cell density. The action of DEX plus EGF was stimulatory but not synergistic in young confluent cultures. DEX plus EGF had no synergistic effect on senescent cells either during log phase or at confluence. Analysis of the effect of DEX on [125I]EGF binding revealed no statistically significant changes in either the number of binding sites or the apparent dissociation constant of the EGF-receptor complex.
Cell Division↗
Inhibition of epidermal growth factor-induced phosphorylation by trifluoperazine.
We studied the effects of a series of drugs on A431, a cell line with well-characterized growth factor requirements and epidermal growth factor (EGF) receptors. The major [32PO4]-labeled protein immunoprecipitated with anti-phosphotyrosine antibodies from EGF-stimulated A431 cells was the EGF receptor. Both the quantity of [32PO4]-labeled EGF receptor immunoprecipitated and the phosphotyrosine content of total [32PO4]-labeled proteins were reduced by the addition during EGF stimulation of trifluoperazine (TFP). TFP had little effect on the binding, internalization, and processing of [125I]-EGF. In addition to the effects on phosphorylation, TFP inhibited cell growth both in the presence and absence of EGF. Morphologically, TFP blocked EGF-induced ruffling. TFP did not alter the EGF-stimulated phosphatidylinositol turnover. In an in vitro experiment using A431 cell membranes, TFP did not inhibit phosphorylation of the EGF receptor.
Animals↗
Cellular senescence: factors modulating cell proliferation in vitro.
In our view, two major areas of the investigation of the aging process have been most fruitful over the past few years: namely, the genetic and hormonal strategies aimed at the understanding of in vitro cellular senescence. The genetic studies have primarily utilized cell fusion techniques and viral probes. Along with cell cycle studies involving the induction of thymidine kinase activity and TTP synthesis, the cell fusion studies are most consistent with a late G1 block in senescent cells. This effect would appear to be distinct from the G0 arrest of density-inhibited or mitogen-restricted cell populations. The hormonal studies which have centered on the regulation of cell proliferation have recently focused on peptide hormones. EGF has been of particular interest since it is so well characterized. This receptor system remains largely unchanged throughout the lifespan with the notable exception of the purified receptor-associated, autocatalytic, tyrosine-specific kinase activity, which decreases with age. The functional significance of this decrease in enzyme activity is unknown, although its growth regulatory importance is implicated in several systems, and may well represent a critical early G0/G1 event which is absent in senescent cells.
Aging↗
The effect of subcutaneous nerve stimulation (SCNS) on pain associated with osteoarthritis of the hip.
A novel form of subcutaneous nerve stimulation (SCNS) was recently introduced for the relief of chronic pain. We present a study using this form of SCNS applied over the radial, median and saphenous nerves in patients with clinically diagnosed osteoarthritis of the hip. Acceptable pain relief was obtained in 60% of patients receiving stimulation, however, comparable analgesia was achieved in a control group, who received no electrical stimulation through similarly placed needles. We suggest that these results may be explained by the ability of SCNS to evoke a placebo response. The efficacy of the placebo effect and the ethical implications of its use in clinical practice are discussed.
Adult↗
Progressive loss of the proliferative response of senescing WI-38 cells to platelet-derived growth factor, epidermal growth factor, insulin, transferrin, and dexamethasone.
Normal human diploid cell lines such as WI-38 display a progressive loss of responsiveness to the mitogenic components in serum. Using a serum-free, hormone-growth factor supplemented medium for WI-38 cells (Phillips & Cristofalo, 1980, 1981), we examined the dose-response relationships of cells to the individual growth factor at various in vitro ages. Although as cultures senesced, they became progressively less responsive to mitogenic stimulation, the concentration of epidermal growth factor (25 ng/ml), insulin (5 micrograms/ml), transferrin (5 micrograms/ml), and dexamethasone (55 ng/ml) that elicited the maximum proliferative response did not change as a function of age. As cultures age they may require increasing amounts of platelet-derived growth factor to elicit the maximum mitogenic response.
Cell Division↗
[125I]EGF binding ability is stable throughout the replicative life-span of WI-38 cells.
Using a serum-free medium supplemented with hormones and growth factors, which included epidermal growth factor (EGF), we investigated the binding and processing-degradation of [125I]EGF in WI-38 cells of various in vitro ages. The binding and processing-degradation systems of these cells remained essentially unchanged throughout their lifespan. The number of specific [125I]EGF binding sites per cell increased as the cultures senesced, though the number of specific binding sites per micron 2 (surface area) remained constant. The kinetics of ligand degradation as well as the qualitative and quantitative nature of the degradation products also remained essentially unchanged throughout the life-span. The only consistent alteration in any of the binding parameters measured was the slight decrease in the apparent Kd of the ligand-receptor complex, independent of temperature. Quantitation of EGF-stimulated DNA synthesis revealed a decrease in the percentage of cells incorporating [3H]thymidine ([3H]TdR) during a 30-h exposure from 45% in young cells to 0.25% in senescent cells, although [125I]EGF binding or processing-degradation did not differ significantly in young and old cells. Thus, EGF binding does not decrease in senescence.
Cell Survival↗
Extradural versus intramuscular diamorphine. A controlled study of analgesic and adverse effects in the postoperative period.
The effects of diamorphine hydrochloride 0.1 mg/kg, given either extradurally or intramuscularly for postoperative analgesia were compared in two randomised double-blind studies involving 39 patients undergoing thoracotomy and major gynaecological surgery. Assessments were made at fixed intervals after the administration of diamorphine and consisted of the measurement of pain or analgesic effect. Segmental, sympathetic and any adverse effects were sought. There was no significant difference in the quality of analgesia between the two groups in either trial. Extradural diamorphine provided safe and effective analgesia of rapid onset, with no specific undesirable side-effects. In both studies, analgesia was more prolonged following extradural administration. The relative proportion of spinal binding may be increased after extradural administration and this may be reflected in the prolonged analgesia observed.
Adult↗
Epidural analgesia following thoracic surgery. A review of two years' experience.
The use of thoracic epidurals for postoperative pain relief in 58 patients following thoracic surgery is reviewed. Epidural catheters were inserted at the end of the operative procedure and it was planned that analgesia was to be maintained with a continuous infusion of bupivacaine. In those patients with evidence of successful neurological blockade there was a high incidence of adverse effects. Hypotension occurred in nearly 80% of patients and inadequate analgesia necessitating intramuscular opiates in 30%. The results suggest that the method adopted is unsuitable for some patients who have undergone major pulmonary surgery, although it has proved a useful adjuvant for pain relief in patients who have had other intrathoracic operations.
Adolescent↗
Histochemical assessment of nuclear protein content and enzyme activity in confluent nonarrested and low-serum arrested WI-38 fibroblasts at mid and late population doubling levels.
Cultures of WI-38 fibroblasts maintained in low-serum medium have little mitotic activity. Such arrested cultures can serve as a useful adjunct to the normal-serum proliferating fibroblast model system of senescence since the reduced cycling activity more accurately reflects in vivo conditions. Normal-serum plateau phase cultures and 17-day low-serum arrested plateau phase cultures were examined at both mid and late population doubling levels (PDL). Acidic, basic and total nuclear protein contents were determined by staining with toluidine blue O, alkaline fast green and acid fast green, respectively. Succinate dehydrogenase and glucose-6-phosphate dehydrogenase were localized with the nitroblue tetrazolium staining reaction. Quantification of individual cell staining was done on a Vickers M85 microdensitometer. Low-serum arrested cells at both mid and late PDL showed a significant increase in acidic protein staining and a decrease in basic protein staining when compared with normal-serum nonarrested cells. Enzyme activities in arrested cells were lower than nonarrested cells at mid PDL but equivalent to nonarrested cells at late PDL. This study demonstrates that the use of selected quantitative histochemical staining in combination with cytophotometry can be employed to differentiate arrested from nonarrested fibroblast populations as well as mid PDL from late PDL cultures.
Blood↗
Hydrocortisone stimulation of DNA synthesis in bromodeoxyuridine-selected non-dividing WI-38 cells.
Ostensibly noncycling WI-38 cells were selected by incubating growing cultures for 7 days with 10(-5) M bromodeoxyuridine. These cells were then exposed to Hoechst dye 33258, and then visible light which kills cells that incorporated bromodeoxyuridine. Following selection, cultures were refed with medium containing 10% fetal bovine serum. By autoradiography, we determined that less than 10% of these cells could incorporate [3H]thymidine during the next 7 days. This value was increased to 25% or more by adding hydrocortisone to the medium. The proliferative response was also increased by refeeding cultures with hydrocortisone-containing medium conditioned 24 h by freshly seeded mid-, but not late, population doubling level (PDL) cultures. Medium conditioned without hydrocortisone did not stimulate incorporation of [3H]thymidine beyond the control value. These results show that: (1) cells that might otherwise not initiate DNA synthesis are stimulated to do so by hydrocortisone; (2) hydrocortisone-conditioned medium from mid-PDL culture increases this stimulation; and (3) hydrocortisone-conditioned medium from late PDL cultures is not stimulatory.
Bromodeoxyuridine↗
Growth regulation of WI38 cells in s serum-free medium.
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Blood↗