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Biomedical subjects

G Brooks

Publications and source records attributed to G Brooks.

At least 55 records · Page 3Linked to original sources
PubMed1994

The role of 80K/MARCKS, a specific substrate of protein kinase C, in cell growth and tumour progression.

Since its discovery more than a decade ago [Wu et al., 1982; Rozengurt et al., 1983], the 80-87 kDa myristoylated alpha lanine-rich C-kinase substrate (80K/MARCKS) protein has attracted a great deal of attention from researchers interested in cell growth and tumour progression. However, despite its ubiquitous distribution, a definitive functional role for 80K/MARCKS has not been found. The purpose of this review is to describe the properties, distribution and regulation of 80K/MARCKS and to discuss some of the most recent findings, both from our laboratory and from others, that have suggested a functional role for this protein in modulating cell growth and tumour progression. Furthermore, I will present data from our laboratory that implicates 80K/MARCKS as a novel tumour suppressor in cells of melanocyte origin.

Actins↗
PubMed1994

Mothers of chronically ill neonates and primary nurses in the NICU: transfer of care.

The purpose of this study was to explore the relationship between nurse and mother during the ill neonate's hospitalization and examine how this relationship influenced the mother's parenting of her infant during the hospitalization. Using qualitative methods, we separately interviewed ten mothers and nine primary nurses about their relationships, their views on each other, and the mothers' infant care. The tape-recorded interviews were transcribed verbatim, themes were extracted, and categories were developed for coding the data. As issues emerged, they were further explored in follow-up interviews. We identified four stages in the process of transferring the care of the infant from nurse to mother. The mother-nurse relationships were influenced by both the nurses' and the mothers' typical interactional patterns. Nurses had definite ideas about who was an "ideal" mother. To the mothers, the nurses' competence and caring attitude toward their infants were most important. The process by which the nurse and mother interact to provide care for the infant and alter their roles during the course of the hospitalization is linked to various factors and aspects of the nurse-mother relationship.

Adult↗
PubMed1993

Growth of melanocytic cells is associated with down-regulation of protein kinase C alpha, delta, and epsilon isoforms. Possible role of diacylglycerol.

Protein kinase C (PKC) down-regulation has been shown to correlate with the growth of murine melanocytic cells in culture (Brooks, G., Wilson, R. E., Dooley, T. P., Goss, M. W., and Hart, I. R. (1991) Cancer Res. 51, 3281-3288). We now show that PKC alpha, delta, epsilon, and zeta isoforms are present at the protein level in quiescent, non-transformed Mel-ab melanocytes, maintained in the absence of phorbol ester. Proliferation of Mel-ab cells, achieved by incubation in the continual presence of phorbol 12,13-dibutyrate, was associated with a down-regulation of the PKC alpha, delta, and epsilon isozymes. Examination of two transformed syngeneic lines (the B16 murine melanoma and the long terminal repeat Ras.2 line), that grew in the absence of exogenous phorbol esters, showed that PKC alpha protein levels were either partially down-regulated or unaffected, the PKC delta and epsilon isoforms were down-regulated completely, and the levels of PKC zeta protein remained unaltered relative to quiescent Mel-ab cells. Basal levels of total diacylglycerol were elevated 5-fold in B16 melanoma cells compared with levels found in quiescent or proliferating Mel-ab melanocytes and appear to arise largely from the breakdown of phosphatidylinositol phospholipids accompanied by a significant rise in phospholipase C activity. Hourly treatments of quiescent Mel-ab melanocytes with the synthetic diacylglycerol analogue, 1,2-dioctanoyl-sn-glycerol, for 24 h, resulted in an induction of DNA synthesis which was associated with a significant down-regulation of PKC levels mediated largely via post-translational rather than transcriptional mechanisms. These results show for the first time that specific isoforms of PKC are down-regulated at the protein level during proliferation of murine melanocytic cells and suggest that the constitutive down-regulation of PKC in transformed melanoma cells may arise as a consequence of elevated endogenous phosphatidylinositol-derived diacylglycerol levels.

Animals↗
PubMed1992

Differential induction of 12-O-tetradecanoylphorbol-13-acetate sequence gene expression in murine melanocytes and melanoma cells.

We previously showed that growth of the nontumorigenic, immortal murine melanocyte line Mel-ab correlates with the depletion of protein kinase C (PKC), whereas quiescence is associated with elevated levels of this enzyme (Brooks G, et al., Cancer Res 51: 3281-3288, 1991). Here we report responses that occur in these cells downstream of PKC activation or downregulation. We examined induction of 12-O-tetradecanoylphorbol-13-acetate (TPA)-inducible sequence (TIS) gene expression in Mel-ab melanocytes and in their transformed counterparts, B16 melanoma cells. Exposure of quiescent Mel-ab cells to the PKC-activating phorbol esters TPA or sapintoxin A at 81 nM for 2 h increased levels of mRNA for six of seven TIS genes examined (twofold to 80-fold increase in steady-state RNA levels for TIS 1, 7, 8, 11, 21, and 28 (c-fos); TIS 10 expression was not affected). No induction of TIS gene expression was observed either in growing Mel-ab cells maintained in 324 nM phorbol 12,13-dibutyrate or in B16 cells previously unexposed to phorbol esters, in which normal PKC levels were endogenously depressed. The cAMP-elevating agents choleratoxin (10 nM) and dibutyryl cyclic AMP (2.5 mM) increased levels of TIS mRNA (with the exception of TIS 10) in both proliferating Mel-ab and B16 cells, suggesting that downregulation of the PKC pathway is specific and not a consequence of a general inhibition of all signalling pathways.

3T3 Cells↗
PubMed1991

Protein kinase C down-regulation, and not transient activation, correlates with melanocyte growth.

The nontumorigenic, immortal line of murine melanocytes, Mel-ab, requires the continual presence of biologically active phorbol esters for growth (R.E. Wilson et al., Cancer Res., 49:711-716, 1989). Comparable treatments of B16 murine melanoma cells result in partial inhibition of cell proliferation. The role of protein kinase C (PKC) in the modulation of growth of cells from these two melanocytic cell lines has been investigated. Significant levels of PKC were present in quiescent Mel-ab cells as determined by Western blotting, whereas no immunoreactive protein was detected in cell extracts from either proliferating Mel-ab or B16.F1 cells. Phosphorylation of a Mr 80,000 protein, which by one- and two-dimensional gel analysis comigrated with the known Mr 80,000 protein substrate of PKC in fibroblasts, was induced in 12-O-tetradecanoylphorbol-13-acetate-stimulated quiescent Mel-ab cells but not in proliferating Mel-ab cells or B16.F1 melanoma cells. Direct measurement of PKC activity in these cells demonstrated a 10-fold greater level of activity in quiescent Mel-ab cells (262 +/- 50 pmol/min/mg SD) compared with growing cells (22.8 +/- 11.8 pmol/min/mg SD). An intermediate level of activity was detected in proliferating B16.F1 melanoma cells (148.5 +/- 20.4 pmol/min/mg SD). The subcellular distribution of PKC was dependent upon the growth state of the cells such that quiescent Mel-ab cells displayed a higher level of activity in the cytosol, whereas growing Mel-ab cells displayed greater activity in the particulate fraction. Like many other transformed lines, B16.F1 melanoma cells constitutively expressed the majority of enzyme activity in the particulate fraction. Measurement of [3H]phorbol ester binding in intact cells paralleled the PKC activation data such that quiescent Mel-ab cells displayed binding of 1612 +/- 147 cpm/10(6) cells, whereas proliferating Mel-ab and B16.F1 melanoma cells displayed binding of 652 +/- 28 and 947 +/- 81 cpm/10(6) cells, respectively. Membrane-permeant diacylglycerol analogues, which activated but did not down-regulate PKC, were devoid of growth-stimulating effects on melanocytes, even in the presence of the specific diacylglycerol kinase inhibitor, R59022. Together, these data show that PKC down-regulation, and not activation, correlates with the growth of melanocytes in culture.

Animals↗
PubMed1991

6-(substituted methylene)penems, potent broad spectrum inhibitors of bacterial beta-lactamase. V. Chiral 1,2,3-triazolyl derivatives.

Structure-activity relationships in a series of (5R)-6-triazolylmethylene penems with potent beta-lactamase inhibitory activity are described. In most cases, their in vitro synergistic activity with amoxycillin is superior to that of clavulanic acid, sulbactam and tazobactam (YTR 830). Against an Escherichia coli TEM-1 infection in mice, the compounds showed a broad range of potencies; an optimum polarity was found, however, which gave maximum potency.

Amoxicillin↗
PubMed1990

Non-promoting diterpene esters can induce Epstein-Barr virus early antigen expression in the Raji cell line.

A range of diterpene ester ligands with selective biological activity (e.g., irritant but not tumour promoting) were tested for their ability to induce Epstein-Barr virus (EBV) early antigen expression in the lymphoblastoid Raji cell line. All C12/C13 substituted compounds were found to be capable of inducing some antigen expression at nM-microM levels, including desacetyl-alpha-sapinine, a compound largely devoid of biological activity. The non-promoting, fluorescent compound, sapintoxin A, was virtually equipotent with promoting compounds. It was concluded that, although the assay has relevance to the specific condition of chronic diterpene ester exposure occurring in conjunction with high EBV infection rates, there was relatively poor correlation with mouse skin tumour promoting potential.

Antigens, Viral↗
PubMed1990

Both tumour-promoting and non-promoting phorbol esters inhibit [125I]EGF binding and stimulate the phosphorylation of an 80 kd protein kinase C substrate protein in intact quiescent swiss 3T3 cells.

Sapintoxin A (SAP A) and 12-deoxyphorbol 13-phenylacetate (DOPP), are two biologically active but non-tumour-promoting phorbol esters that potently bind to and activate the phorbol ester receptor, protein kinase C (PKC). SAP A and DOPP cause a dose-dependent increase in the phosphorylation of an 80 kd (80K) substrate protein for PKC in Swiss 3T3 cells. A similar dose-response effect was seen with sapintoxin D (SAP D), the stage 2 promoting analogue of 12-O-tetradecanoylphorbol-13-acetate and the complete promoter phorbol 12,13-dibutyrate (PDB). The doses resulting in a half maximal phosphorylation of this protein (Ka) were 20 nM (SAP A), 45 nM (DOPP), 23 nM (SAP D) and 37 nM (PDB). Both non-promoting and promoting phorbol esters induced a dose-dependent inhibition of [125I]epidermal growth factor (EGF) binding to its receptor in Swiss 3T3 cells. The doses required for 50% inhibition of binding (Ki) were: 8 nM (SAP A), 16 nM (DOPP), 14 nM (SAP D) and 17 nM (PDB). The results clearly demonstrate that induction of phosphorylation of the 80K phosphoprotein and inhibition of [125I]EGF binding in Swiss 3T3 cells following exposure to phorbol esters is independent of the tumour-promoting activity of these compounds. The fact that SAP A, DOPP, SAP D and PDB are mitogenic for a variety of cell types and that exposure to these compounds leads to 80K phosphorylation and inhibition of [125I]EGF binding, suggests that these early biological events may play a role in the mitogenic response induced by these compounds.

Animals↗
PubMed1990

Inhibition of [125I]epidermal growth factor binding to murine fibroblasts and keratinocytes by irradiation with UV-B light. Evidence for a protein kinase C-independent mechanism.

Treatment of murine Swiss 3T3 fibroblasts and XB/2 keratinocytes with UV-B light (302 nm) resulted in a dose-dependent inhibition of [125I]epidermal growth factor (EGF) binding. The light dose required to achieve 50% inhibition of binding in both cell types was 80-85 J/m2. Decreased [125I]platelet-derived growth factor binding was not evoked even by light doses of up to 280 J/m2. UV-B irradiation did not stimulate phosphorylation of the 80 kd protein substrate for protein kinase C. Furthermore, its effect on [125I]EGF binding was not altered as a consequence of protein kinase C down-regulation following prolonged exposure of cells to phorbol esters. These results indicate that UV-B-induced transmodulation of the epidermal growth factor receptor is a specific event mediated through a protein kinase C-independent pathway. Transfer of culture medium from irradiated cells to untreated control cells showed this effect was not induced as a result of transforming growth factor alpha release and subsequent binding to the EGF receptor in these cells.

Animals↗
PubMed1990

Maxillofacial fractures in the elderly: a comparative study.

Previous maxillofacial trauma research has dealt primarily with facial bone fractures in the general population. Very few studies have specifically addressed maxillofacial fractures in the elderly. We compared 45 elderly (65 years of age or older) and 201 younger adult (16 to 64 years of age) patients admitted to our hospital with maxillofacial fractures. The percentage of patients admitted with nasal bone fractures was much greater in the elderly population, while mandibular fractures were more common in the adult group. Motor vehicle accidents accounted for over half the injuries in both groups, while falls were more prevalent in the elderly. Management of the elderly patient may be complicated by their associated injuries or underlying medical problems, perhaps partially accounting for their longer median length of hospital stay. The elderly are a unique subpopulation of maxillofacial fracture patients and deserve further study regarding their injuries and optimal methods for treatment.

Accidental Falls↗
PubMed1989

Tumour-promoting and hyperplastic effects of phorbol and daphnane esters in CD-1 mouse skin and a synergistic effect of calcium ionophore with the non-promoting activator of protein kinase C, sapintoxin A.

Using an 18 week two-stage protocol we have compared the tumour-promoting properties of a range of phorbol and daphnane esters on female CD-1 mice. The induction of epidermal hyperplasia in this mouse strain by these compounds has also been assessed by comparison with the standard phorbol ester, 12-O-tetradecanoylphorbol-13-O-acetate (TPA). Two compounds, sapintoxin D (SAP D) and thymeleatoxin A (TA) (a daphnane structurally related to the second-stage promoter mezerein) were shown to be second-stage promoters using 5 nmol TPA as a first-stage promoter and 0.2 mumol 7,12-dimethylbenz[a]anthracene (DMBA) as initiator. Both compounds at a dose of 17 nmol were hyperplasiogenic. Two further derivatives, sapintoxin C (SAP C) and 4 alpha-sapinine (alpha-SAP) were inactive as promoters and hyperplastic agents. 4 alpha-sapinine, which prevents in vitro stimulation of protein kinase C (PKC), by 12-O-tetradecanoylphorbol-13-O-acetate (TPA) failed to inhibit significantly TPA-induced promotion and hyperplasia at a dose of 20 nmol and 100 nmol respectively. Sapintoxin A (SAP A), a potent activator of PKC, was neither a complete nor second-stage promoter at doses of up to 20 nmol. A series of in vivo and in vitro experiments which were carried out to determine the metabolic fate of this compound under experimental conditions showed that SAP A was not metabolized to any significant extent up to 48 h. When SAP A was co-administered with sub-hyperplastic doses of the calcium ionophore A23187 (5 micrograms and 10 micrograms) tumours appeared in a dose-dependent manner. This combination was also hyperplasiogenic in mouse skin. SAP A may be a useful probe for studying the involvement of PKC isozymes in tumour promotion and cell proliferation.

Animals↗
PubMed1987

Toxic phorbol esters from Chinese tallow stimulate protein kinase C.

Phorbol esters were isolated from the seeds of Chinese tallow (Sapium sebiferum L. Roxb.). These compounds were based on the tigliane nuclei, 4-deoxyphorbol, 12-deoxyphorbol and 4,20-dideoxy-5-hydroxyphorbol. The pro-inflammatory activity (ID50) of the pure compounds was between 0.042 and 2.6 nmoles per ear. Protein kinase C activation assays were carried out on samples of enzyme purified from mammalian brain and the activities (Ka) were in the range 76-176 nM. The 4,20-dideoxy-5-hydroxy analogue was inactive in both tests. Chinese tallow, which is used as a substitute for linseed oil, may represent an industrial toxic hazard in terms of both pro-inflammatory and tumour-promoting effects.

Animals↗
PubMed1987

Sapintoxin A. A fluorescent phorbol ester that is a potent activator of protein kinase C but is not a tumour promoter.

In this communication we report on the activity of the naturally occurring, highly fluorescent phorbol ester Sapintoxin A (12-o-[2-methylaminobenzoate]-4-deoxyphorbol 13-acetate). This compound potently activates the enzyme protein kinase C (PKC) (Ka 76 nM) but is neither a complete nor second-stage tumour promoter in traditional Berenblum tests. Sapintoxin A has properties in common with promoters such as 12-o-tetradecanoylphorbol 13-acetate (TPA) in that it will induce erythema in vivo, induce lymphocyte mitogenesis in vitro and cause aggregation of human and rabbit platelets. Accordingly, Sapintoxin A is a suitable negative control compound for biochemical studies concerning the involvement of PKC in tumour promotion and cell proliferation.

Animals↗