Amount Of Platelet-derived Growth Factor Research Articles

The effects of platelet-rich plasma on the proliferation and release of growth factors from periodontal ligament cells

Periodontal ligament cells (PDLCs) play an important role in the regeneration of periodontium. The healing potential of PDLCs may be due to the high concentration of growth factors in platelet-rich plasma (PRP). In this study, the effects of PRP on the proliferation and activation of PDLCs and growth factor release from PDLCs were investigated. PDLCs were isolated from third molars or premolars of healthy patients. Whole blood was obtained from healthy volunteers for the preparation of activated PRP. The platelet concentration in PRP was measured and the amount of platelet-derived growth factor (PDGF)-AB, PDGF-BB, transforming growth factor-β1 (TGF-β1), and vascular endothelial growth factor (VEGF) were determined by enzyme-linked immunosorbent assay. The effects of activated PRP on PDLC proliferation, attachment, alkaline phosphatase (ALP) activity, and growth factor release were investigated. Platelet concentration was increased 5.41-fold in PRP compared to whole blood. PDGF-AB, PDGF-BB, TGF-β1, and VEGF were detected in PRP at concentrations of 273.38 ng/mL, 47.0 ng/mL, 168.42 ng/mL, and 510.56 pg/mL, respectively. PDLCs cultured with ≥10% PRP showed significantly increased cell proliferation and ALP activity compared to the control (p<0.05). PDLCs cultured with 10% PRP also presented higher cell attachment and increased release of TGF-β1 and VEGF compared to the control (p<0.05). PRP can deliver high concentrations of growth factors to a defect site to increase the proliferation, attachment, and ALP activity of PDLCs. These results suggest that PRP might effectively contribute to periodontal regeneration.

AB84. Characteristic of platelet-rich plasma and its effects on on improving neurogenic erectile dysfunction

Platelet-rich plasma (PRP) containing autologous thrombocyte growth factors is applied in regenerative medicine, but the lack of an optimized PRP preparation protocol causes unstable therapeutic effects. The aim of this study was to optimize the PRP preparation method to obtain the largest amount of growth factors released and compare the effects of PRP from different preparation methods in restoring of erectile function in bilateral cavernous nerve (CN) injury rat model. The in vivo experiments used Sprague-Dawley male rats (n=24) which were randomly divided into four groups of equal numbers: (I) sham operation; (II) vehicle only (which underwent bilateral nerve crushing without treatment); (III) general PRP; and (IV) optimized PRP containing the largest amount of platelet-derived growth factor (PDGF)-AB (groups 3 and 4 underwent bilateral CN crushing with an immediate injection of the respective PRP into the corpus cavernosum). The intracavernous pressure (ICP) of all rats was monitored. Results demonstrated that in the PRP group prepared with the anticoagulant, ACD-A, the amount of PDGF-AB released was statistically higher than those of the other groups (P<0.05). Greater concentrations of PDGF-AB were measured in PRP after chitosan treatment compared to treatment with a solvent or other activating factors. Additionally, storing PRP at –20 ℃ resulted in a significant increase in PDGF-AB released compared to storage at other temperatures. The optimized PRP prepared by stimulation with chitosan and incubated at –20 ℃ for 15 days had the largest amount of PDGF-AB and showed a synergistic effect on release (P<0.05). The functional outcome measurement revealed improvement after bilateral CN injury occurred in the group treated with optimized PRP. It was concluded that optimized PRP with a high level of growth factors was more stable, and its injection into the corpus cavernosum facilitated recovery of erectile function.

AB84. Characteristic of platelet-rich plasma and its effects on on improving neurogenic erectile dysfunction

Platelet-rich plasma (PRP) containing autologous thrombocyte growth factors is applied in regenerative medicine, but the lack of an optimized PRP preparation protocol causes unstable therapeutic effects. The aim of this study was to optimize the PRP preparation method to obtain the largest amount of growth factors released and compare the effects of PRP from different preparation methods in restoring of erectile function in bilateral cavernous nerve (CN) injury rat model. The in vivo experiments used Sprague-Dawley male rats (n=24) which were randomly divided into four groups of equal numbers: (I) sham operation; (II) vehicle only (which underwent bilateral nerve crushing without treatment); (III) general PRP; and (IV) optimized PRP containing the largest amount of platelet-derived growth factor (PDGF)-AB (groups 3 and 4 underwent bilateral CN crushing with an immediate injection of the respective PRP into the corpus cavernosum). The intracavernous pressure (ICP) of all rats was monitored. Results demonstrated that in the PRP group prepared with the anticoagulant, ACD-A, the amount of platelet-derived growth factor (PDGF)-AB released was statistically higher than those of the other groups (P<0.05). Greater concentrations of PDGF-AB were measured in PRP after chitosan treatment compared to treatment with a solvent or other activating factors. Additionally, storing PRP at –20 °C resulted in a significant increase in PDGF-AB released compared to storage at other temperatures. The optimized PRP prepared by stimulation with chitosan and incubated at -20 °C for 15 days had the largest amount of PDGF-AB and showed a synergistic effect on release (P<0.05). The functional outcome measurement revealed improvement after bilateral CN injury occurred in the group treated with optimized PRP. It was concluded that optimized PRP with a high level of growth factors was more stable, and its injection into the corpus cavernosum facilitated recovery of erectile function.

Optimization of platelet-rich plasma and its effects on the recovery of erectile function after bilateral cavernous nerve injury in a rat model

Platelet-rich plasma (PRP) containing autologous growth factors is applied in regenerative medicine, but the lack of an optimized PRP preparation protocol causes unstable therapeutic effects. The aim of this study was to optimize the PRP preparation method and compare the effects of PRP from different preparation methods in restoration of erectile function in a rat model. The in vivo experiments used Sprague-Dawley male rats (n = 24), which were randomly divided into four groups of equal numbers: group I underwent sham operation, while the remaining three groups underwent bilateral CN crush. Crush injury groups were treated at the time of injury with an application of general PRP, optimized PRP [with the largest amount of platelet-derived growth factor (PDGF)-AB] or normal saline-only injection in the corpus cavernosum, respectively. Four weeks later, erectile function was assessed by CN electrosimulation, and penile tissue was collected for histology. Results demonstrated that in the PRP group prepared with the ACD-A anticoagulant, chitosan and incubated at -20°C for 15 days had the largest amount of PDGF-AB and showed a synergistic effect on release (p < 0.05). Functional outcome measurement and immunofluorescence staining for the dorsal nerve revealed that improvement after bilateral CN injury occurred in the optimized PRP group (p < 0.05). It was concluded that optimized PRP with a high level of growth factors was more stable, and its injection into the corpus cavernosum facilitated recovery of erectile function. Copyright © 2013 John Wiley & Sons, Ltd.

Effect of Nitric Oxide Donor on IL-6 and PDGF-BB: Comparison of Diabetic Versus Normal Tissue Under Hypoxic and Normoxic Conditions

Statement of the Problem: CDC estimates that 20.8 million people in the US have diabetes. Vascular changes in diabetic tissues result in decreases in oxygenation, platelet derived growth factor (PDGF), and nitric oxide (NO). Fibroblasts and collagen production are significantly reduced in cells from diabetic patients1. Decreased NO production and increased matrix metalloproteinase 8 and 9 occur in wounds of diabetic patients2. Diabetic wounds demonstrate increased inflammatory cytokine secretion, including Interleukin-6. This lab has also demonstrated that the addition of an NO donor can reduce the MMP-8 and MMP-9 expression by fibroblastic cells3. Materials and Methods: Human skin fibroblasts from a diabetic subject were compared to fibroblasts from a non-diabetic patient grown under normoxic and hypoxic conditions. IL-6 and PDGF production were measured. Fibroblasts from a 31yo diabetic and 32yo non-diabetic patients were obtained from the Coriell Institute. Fibroblasts were cultured to confluence and treated with 1nM, 10nM, or 100nM of long-acting SNitrosoglutathione(SNOG) or short-acting FK409 (NOR3) NO donor for 1, 3, or 7 days. Cells were cultured under normoxic (20% oxygen) or hypoxic (2% oxygen) conditions. Appropriate untreated controls were included in all experiments. At harvest, conditioned media from the cultures were assayed for production of IL-6 or PDGF-BB. IL-6 was measured using a ChemiKine® Human Interleukin-6 Sandwich ELISA kit (Chemicon International). PDGF-BB was measured using a Quantikine® Human Immunoassay (RD where significant differences were found, a two-tailed t-test was performed. P-values less than 0.05 were considered significant. Results: After 1, 3, and 7 days in culture, normal fibroblasts produced 23% as much IL-6 as diabetic cells under normoxic conditions. Under hypoxic conditions, normal cells produced an even smaller relative amount of IL-6 (14%). The production of IL-6 by diabetic, but not normal, fibroblasts increased with time in culture. Under normoxic conditions, IL-6 production by diabetic fibroblasts increased from 4.34ng/mg protein to 6.94ng/mg protein from day 1 to day 7, under hypoxic conditions, IL-6 increased from 11.23ng/mg protein to 17.98ng/mg protein. NOR3 had no effect on IL-6 production by any cells under either condition. Treatment with SNOG, decreased IL-6 production by both normal and diabetic fibroblasts, with greater effect on diabetic fibroblasts. Under normoxic conditions, IL-6 production by diabetic cells was dosedependently decreased by up to 81% at the 100nM dose. A smaller response was observed with normal fibroblasts. Only the 100nM dose resulted in a significant decrease in IL-6 production at all three time points. Cells cultured under hypoxic conditions had similar results, but the magnitude was greater in diabetic cells. PDGF-BB was produced by both normal and diabetic fibroblasts. Normal fibroblasts under normoxic conditions synthesized twice as much PDGF-BB as diabetic cells. Under hypoxic conditions, normal cells produced five times the amount of PDGF synthesized by diabetic cells. There were no significant changes in PDGF-BB production with NO donor treatment. Conclusions: Production of PDGF-BB, is significantly reduced under hypoxic conditions in both normal and diabetic cells, but the effect is much greater in the diabetic cells. Synthesis of PDGF-BB by diabetic cells is more influenced by hypoxia than normal cells. Treatment with the NO donor compounds had no effect on PDGF-BB production. Production of IL-6, an inflammatory mediator, is increased in diabetic cells under both normoxic and hypoxic conditions. The addition of SNOG, significantly decreased the amount of IL-6 produced by normal and diabetic cells, but the effect was greater in the diabetic cells.

Transferrin up-regulates chemokine synthesis by human proximal tubular epithelial cells: Implication on mechanism of tubuloglomerular communication in glomerulopathic proteinura

The pathogenesis of glomerulosclerosis and tubulointerstitial fibrosis in proteinuric renal disease is obscure. We recently showed that transferrin, a key proteinuric component, mediates proximal tubular epithelial cell (PTEC) C3 synthesis. To further examine whether proteinuric tubular injury may induce glomerular inflammation and to characterize the role of transferrin in activating PTEC, glomerular mesangial cells (MC) were exposed to transferrin-activated PTEC culture supernatant and their proliferative and profibrotic responses analyzed. Human PTEC and MC were obtained by primary culture. Confluent, transferrin-stimulated PTEC were grown in serum-free medium to produce a "conditioned" medium that was incubated with quiescent MC. The proliferative response of MC was then assessed by thymidine uptake, and the expression of fibrogenic factors measured by reverse transcription-polymerase chain reaction (RT-PCR) and enzyme-linked immunosorbent assay (ELISA). The chemokine profile in PTEC after transferrin treatment was examined by RT-PCR and ELISA. "Conditioned" supernatant from PTEC, which contained the highest amounts of platelet-derived growth factor (PDGF), stimulated MC proliferation compared with serum-free (P = 0.03) or transferrin-containing (P = 0.009) control media. This proliferative response was partially abrogated by treating MC with anti-PDGF. MC expression of PDGF, but not transforming growth factor-beta or intercellular cell adhesion molecule-1, was up-regulated by conditioned PTEC medium. Transferrin up-regulated monocyte chemoattractant peptide-1, interleukin-8, and macrophage migration inhibitory factor expression in a time- and dose-dependent fashion, but had no effect on RANTES expression by PTEC. These results provide experimental evidence suggesting that there is a tubuloglomerular "cross-talk" mechanism in the proteinuric state. PTEC-secreted PDGF, which further induces mesangial PDGF, could partially account for the mesangial proliferation frequently observed in proteinuric renal disease. Transferrin is one of the culprit nephrotic proteins leading to tubular overexpression of various proinflammatory chemokines, which may explain the interstitial changes observed in proteinuric states.

Studies on the Biological Effects of Ozone: 10. Release of Factors from Ozonated Human Platelets

In a previous work we have shown that heparin, in the presence of ozone (O3), promotes a dose-dependent platelet aggregation, while after Ca2+ chelation with citrate, platelet aggregation is almost negligible. These results led us to think that aggregation may enhance the release of platelet components. We have here shown that indeed significantly higher amount of platelet-derived growth factor (PDGF), transforming growth factor beta1 (TGF-beta1) and interleukin-8 (IL-8) are released in a dose-dependent manner after ozonation of heparinised platelet-rich plasma samples. These findings may explain the enhanced healing of torpid ulcers in patients with chronic limb ischemia treated with O3 autohaemoteraphy (O3-AHT).

Tyrosine phosphorylation of platelet derived growth factor β receptors in coronary artery lesions: implications for vascular remodelling after directional coronary atherectomy and unstable angina pectoris

BackgroundGrowth factors such as platelet derived growth factor (PDGF) have been postulated to be important mediators of neointimal proliferation observed in atherosclerotic plaques and restenotic lesions following coronary interventions. Binding...

Platelet-derived growth factor production by cells from Dacron grafts implanted in a canine model.

Previous studies of grafts implanted in dogs documented a time-dependent increase in platelet-derived growth factor (PDGF) production that correlated with inner-capsule thickness. The purpose of this study was to identify the cells in vascular grafts that produce PDGF. Dacron thoracoabdominal grafts were seeded with autologous endothelial cells (ECs), implanted in 11 beagles, and removed after 4 or 20 weeks. ECs and smooth muscle cells (SMCs) were cultured from grafts and adjacent aorta, and PDGF in the conditioned media was measured by radioreceptor assay. The PDGF A-chain mRNA level in freshly harvested cells was assessed using reverse transcriptase, followed by polymerase chain reaction, and expressed as a ratio of glyceraldehyde-3-phosphate dehydrogenase signal. Localization of PDGF A-chain and B-chain protein was also examined with immunohistochemical analysis. Graft and aortic ECs in primary culture did not produce significantly different amounts of PDGF in 72 hours, averaging 368 +/- 160 and 340 +/- 81 pg/microgram DNA, respectively. Graft SMCs in primary culture produced significantly more PDGF than aortic SMCs (584 +/- 343 and 113 +/- 94 pg/microgram DNA, respectively; p < 0.01). Graft SMC PDGF secretion remained greater than aortic SMC PDGF secretion through at least six cell passages. PDGF A-chain mRNA levels were not significantly different for aortic or graft ECs. The PDGF A-chain mRNA level was significantly higher for graft SMCs than aortic SMCs (2.44 +/- 0.67 and 1.45 +/- 0.57 pg/microgram, respectively; p < 0.03). Immunocytochemical analysis detected PDGF A-chain and B-chain protein in the ECs from both native aorta and graft as well as the subendothelial SMCs in the graft, but not in the SMCs of the native aorta. These results suggest that graft SMCs are functionally altered, producing more PDGF than aortic SMCs. PDGF produced by graft SMCs may contribute to the development of intimal hyperplasia.

Growth factors and canine flexor tendon healing: Initial studies in uninjured and repair models

The role of growth factors in a variety of bone and soft tissue healing processes has been studied extensively in numerous recent models, yet little is known about the specific growth factors that may be playing a role in flexor tendon healing. We used a number of established protein purification techniques and bioassays to isolate and partially characterize a heparin-binding growth factor from unoperated canine tendons. Our data provide evidence that basic fibroblast growth factor, a potent angiogenic growth factor, is present in normal canine intrasynovial flexor tendons. We then studied repaired canine flexor tendons to further elucidate the role of growth factors in the tendon healing process. Heparin-sepharose elution profiles from three repair intervals (3, 10, and 17 days) were graphed and compared to known profiles of isolated growth factors. The three repair intervals demonstrated two elution profile peaks, consistent with varying amounts of platelet-derived growth factor and epidermal growth factor. Although additional experimentation is required to identify definitively the various protein isolates, these data provide compelling evidence that a variety of growth factors are present in uninjured and healing digital flexor tendons.

Presence of Platelet-derived Growth Factor in Normal and Fibrotic Lung Is Specifically Associated with Interstitial Macrophages, While Both Interstitial Macrophages and Alveolar Epithelial Cells Express the c-sis Proto-oncogene

Normal lung structure is maintained by the presence of mesenchymal cells and their extracellular matrix products. The slow normal turnover of these cells is disrupted in fibrotic disorders, resulting in the in situ accumulation of mesenchymal cells and their extracellular matrix leading to a progressive alveolar wall thickening. Idiopathic pulmonary fibrosis (IPF) is a chronic fibrotic disorder of the lung characterized by a diffuse interstitial and intra-alveolar inflammation dominated by macrophages and polymorphonuclear neutrophils. Evaluation of alveolar macrophages (AM) obtained by bronchoalveolar lavage has previously shown that AM from normal individuals spontaneously release small amounts of platelet-derived growth factor (PDGF), a chemotactic and growth factor for mesenchymal cells, whereas AM from IPF patients spontaneously release increased amounts of biologically active PDGF, suggesting its involvement in mesenchymal cell accumulation. However, other cells such as endothelial cells and vascular smooth muscle cells can also release PDGF in vitro. In order to specify PDGF location in lung parenchyma, open lung biopsies from normal individuals and IPF patients were examined by immunohistochemistry using an anti-PDGF antibody and by in situ hybridization using PDGF A-chain and B-chain gene probes. In normal as well as in fibrotic lung, PDGF was only present in relation with interstitial macrophages but not with any other inflammatory cells or mesenchymal cells. Furthermore, the percentage of PDGF-positive macrophages in IPF was 3-fold increased in comparison to normal lung. In addition, the percentage of PDGF-positive macrophages was the same in fibrotic and nonfibrotic areas of IPF lungs.(ABSTRACT TRUNCATED AT 250 WORDS)

Specific growth factor activity identifies and predicts murine mammary tumor

In milk from substrains of mice with varying incidences of developing mammary tumor, we have isolated a specific mitogenic activity, which is unique in that it identifies mice with mammary tumor and predicts those mice that will eventually develop mammary tumor. None of the milk samples from control mice, who never developed mammary tumor, contained this specific predictive mitogenic activity. Chemical characterization has shown this specific mitogenic activity to be acid- and heat-stable and resistant to reducing agents. Partial purification, by ion-exchange, high-performance liquid chromatography size-exclusion, and isoelectrofocusing techniques, of this specific mitogenic activity from milk of mice that had or eventually developed mammary tumor identifies several peptide growth factor components in a 6–10 kDa molecular weight range. Of known growth factors, radioassay techniques identify an insulin-like growth factor-1-like peptide as a major component. Small amounts of platelet-derived growth factor and transforming growth factor-β activities also were present. Our results suggest that a subset of growth factors that are diagnostic of the presence of murine mammary tumor and predictive of eventual tumor development may be early indicators of the transition of a competent cell to a progressively malignant state. Similar studies of a secreted body fluid from women at risk for breast cancer may lead to the identification of a specific biologic tumor marker for breast cancer.

Exaggerated spontaneous release of platelet-derived growth factor by alveolar macrophages from patients with idiopathic pulmonary fibrosis.

Idiopathic pulmonary fibrosis is a fibrotic lung disease characterized by an increased number of mesenchymal cells in the alveolar walls. Alveolar macrophages constitutively express low levels of c-sis, the protooncogene coding for the B chain of platelet-derived growth factor, a protein with chemotactic and mitogenic activity toward mesenchymal cells. We therefore hypothesized that alveolar macrophages in patients with idiopathic pulmonary fibrosis may release increased amounts of platelet-derived growth factor, which might help to explain the accumulation of mesenchymal cells and the fibrosis of the lower respiratory tract in the disease. Evaluation of alveolar macrophages recovered from the lungs of patients with idiopathic pulmonary fibrosis demonstrated that these cells spontaneously released four times more platelet-derived growth factor than did alveolar macrophages recovered from normal persons (P less than 0.01). That the platelet-derived growth factor molecules were potentially active was shown by their chemotactic activity for smooth-muscle cells and their ability to act as a "competence" factor for fibroblast growth. These observations suggest the possibility that the accumulation of mesenchymal cells within the alveolar walls in patients with idiopathic pulmonary fibrosis may result partly from the exaggerated release of the potent mitogen platelet-derived growth factor by mononuclear phagocytes in the lower respiratory tract.

Production of PDGF-like growth factors by embryonal carcinoma cells and binding of PDGF to their endoderm-like differentiated cells

In this report, we demonstrate that F9 and PC-13 embryonal carcinoma (EC) cells do not bind significant amounts of platelet-derived growth factor (PDGF), whereas the endoderm-like differentiated cells derived from EC cells do. The F9-differentiated cells exhibit approximately 8300 receptors per cell, with an apparent dissociation constant of 30 p M. Two endoderm-like cell lines, PSA-5E and PYS-2, also bind PDGF and exhibit approximately 4800 and 23,500 receptors per cell, respectively. The lack of PDGF binding by the parental EC cells is consistent with their release of a factor(s) that is closely related to PDGF. This factor(s) competes with PDGF for binding to membrane receptors and is recognized by antibodies raised against PDGF. However, this factor(s) does not appear to be antigenically identical to PDGF. We also show that production of this PDGF-like factor(s) is reduced more than 90% when F9 EC cells differentiate into cells that bind PDGF. Thus, our findings indicate that EC cells release a factor(s) that should be capable of binding to their differentiated cells. This raises the possibility that PDGF, or a closely related factor, can influence cell proliferation and/or cell behavior of early embryonic cells.

Modulation of the platelet-derived growth factor induced replicative response.

Quiescent cultures of density arrested BALB/c-3T3 cells have been sensitized to the growth stimulatory action of the platelet-derived growth factor (PDGF). Sensitization was achieved by depriving the cultures of PDGF prior to growth stimulation and was noted after transfer of cultures from medium supplemented with 10% serum to medium containing either an equivalent concentration of platelet-poor plasma or a low concentration (0.5%) of serum. Sensitized cultures required less pure PDGF for growth stimulation than nonsensitized ones. In addition such cultures required less mitogen to synthesize a PDGF modulated major excreted protein (MEP). The mechanism of sensitization was investigated. Sensitized cultures did not bind more PDGF than non-sensitized ones. Rather, sensitization appeared to result from the loss of cells that occurred when cultures were deprived of PDGF. Such a loss increased the amount of PDGF available per cell, causing a higher percentage of cells to enter the S phase. Similarly, the amount of PDGF per cell regulated MEP synthesis. Furthermore, in non-sensitized cultures (containing the same number of cells), the absolute quantity rather than the concentration of PDGF regulated DNA synthesis. It appears that the amount of PDGF per cell modulates mitogenesis.

Macrophage-Derived Growth Factor for Fibroblasts and Interleukin-1 Are Distinct Entities

P388D1, a mouse macrophagelike cell line, was adapted to grow continuously in an unsupplemented, serum-free culture medium and continued to elaborate substances that were mitogenic for quiescent mouse fibroblasts (BALB/c 3T3 cells) and for thymocytes suboptimally stimulated with lectins. We have previously described [37] the fibroblast mitogenic activity as a macrophage-derived competence factor (MDCF). Serum-free, macrophage-conditioned culture medium was concentrated 1,000-fold by a combination of ultrafiltration (hollow fiber) and lyophilization. Concentrates of medium were subjected to gel filtration (Sephadex G-75 or G-150), and the fractions were assayed for mitogenic activity (MDCF) on density-arrested BALB/c 3T3 cells and for Interleukin-1 (IL-1) activity in suboptimally stimulated (Con A) mouse thymocytes. The apparent molecular weight (MW) of MDCF activity was estimated at 56,000 daltons, whereas the peak of IL-1 chromatographed at an apparent MW of 14-16K daltons. There was no detectable IL-1 activity in the MDCF fractions and no detectable MDCF in the IL-1 fractions. These data indicate that P388D1 cells produce both MDCF and IL-1 activities under continuous serum-free conditions and that the two activities are not identical. Stimulation of responsive mononuclear phagocytes with lipopolysaccharide and/or lymphokine-rich supernates resulted in a differential modulation of MDCF and IL-1 activities. Finally, antibody-purified IL-1 had no significant ability to stimulate DNA synthesis in quiescent fibroblasts at concentrations that were mitogenic for thymocytes. However, IL-1 did augment the mitogenic activity of suboptimal amounts of platelet-derived growth factor (PDGF), another competence factor. Further studies revealed that neither the generation nor the activity of MDCF was modulated by the presence of various inhibitors of proteolytic enzymes.

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.