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Biomedical subjects

R M Bell

Publications and source records attributed to R M Bell.

At least 37 records · Page 2Linked to original sources
PubMed1997

Lipid biochemistry: functions of glycerolipids and sphingolipids in cellular signaling.

Products of glycerolipid and sphingolipid metabolism are now known to fulfill second messenger functions in a variety of cellular signaling pathways. Evidence for glycerolipid-derived second messengers was first obtained from the "phosphatidylinositol cycle," which involves a signal-dependent hydrolysis of phosphatidylinositol bisphosphate yielding diacylglycerol and inositol trisphosphate. The role of diacylglycerol in the regulation of protein kinase C activity and its site of interaction with PKC are now well known. Recently, another glycerolipid second messenger, phosphatidic acid, was found to interact with the protooncogenic Raf-1 kinase. In cultured cells, a signal-induced generation of phosphatidic acid was critical for Raf-1 translocation to the cell membrane. Thus, different glycerolipid second messengers appear to regulate distinct targets with exquisite specificity. Analogous to the PI cycle, a "sphingomyelin cycle" was also found to exist, generating sphingolipid second messengers. Ceramide, derived from the agonist-induced hydrolysis of sphingomyelin, is a potent biomolecule with effects in multiple cell signaling pathways. The steroid hormone progesterone stimulated sphingomyelin hydrolysis in Xenopus oocytes. Ceramide, derived from the "sphingomyelin cycle," was sufficient for meiotic cell cycle progression in the oocytes. These results demonstrate the various effects of lipid-derived second messengers and promise exciting discoveries into the roles of lipids in cell signaling.

Animals↗
PubMed1997

Students' acquisition and use of school condoms in a high school condom availability program.

OBJECTIVES: To determine what students know about a condom availability program in their high school, how they react to the program, whether they obtain condoms from it, and what they do with these condoms. DESIGN: Self-administered anonymous survey conducted 1 year after the program began. SETTING: An urban California school district. PARTICIPANTS: A total of 1112 students, 9th through 12th grade, 59% of eligible students present on the survey day. MAIN OUTCOME MEASURES: History of obtaining condoms from the program, use of these condoms, knowledge about the program, and attitudes toward the program. RESULTS: Forty-eight percent of students had personally taken school condoms, and another 5% had gotten them from someone else, for a total of 53% who had obtained school condoms. Seventy percent of nonvirgins and 38% of virgins obtained condoms. Males were more likely than females to have obtained condoms (60% vs 45%). Fifty-four percent of students who had obtained school condoms had used them for sexual activity: 52% had used them for vaginal intercourse, 7% for anal intercourse, and 4% for fellatio. Students also explored school condoms without having sex, eg, removing them from the packet, putting them on fingers, or putting them on their penis or a partner's penis. Thirty-four percent of students who had used a condom for vaginal intercourse during the previous year had obtained the condom they last used from school, with more males than females reporting the school as their source (41% vs 26%). Eighty-eight percent of students knew that all students were allowed to obtain condoms, and 74% knew that parental permission was not required. Students generally supported the condom program: 88% thought the school should give out condoms, and 79% thought that if the school were to require parental permission for students to get condoms, students would get them less often than with the present system (which does not require permission). Thirteen percent agreed and 71% disagreed that "having condoms available at school makes it harder for someone who doesn't want to have sex to say no." CONCLUSIONS: Providing high school students with direct access to condoms leads to widespread use of school condoms, both for sexual activity and for exploratory activities that familiarize students with condoms. Condoms are of interest to both students who have and students who have not engaged in sexual activities for which condoms are recommended.

Adolescent↗
PubMed1997

Adjusting cesarean delivery rates for case mix.

OBJECTIVES: (1) To describe the issues in developing a clinical predictor of cesarean delivery that could be used to adjust reported cesarean rates for case mix, and (2) to compare its performance to other, simpler predictors using clinical and statistical criteria. DATA SOURCES: Singleton births greater than 2,500 grams in Washington State in 1989 and 1990 for whom mothers and infant hospital discharge records could be matched to birth certificate data. DESIGN: Statistical analysis of retrospective merged hospital and birth certificate data, which were used to develop variables and models to predict the probability that any particular delivery would be a cesarean. PRINCIPAL FINDINGS: Merged data led to better predictor variables than those based on one source. A simple four-category hierarchical classification into births with prior cesarean, breech but no prior cesarean, first birth, and other explains 30 percent of the variance in individual cesarean rates. The full clinical model fit the data well and explained 37 percent of the variance. Multiparas without serious complications comprised 35 percent of the mothers and averaged less than 2 percent cesareans. A hospital's predicted cesarean rate depends strongly on the proportion of its births that are first births. CONCLUSION: Government and private agencies have reported cesarean rates as measures of hospital performance. Depending on data and resources available, both simple and complex measures of case mix can be used to adjust reported rates. These adjustments should not include all variables related to the rates. Proper adjustments may not alter hospital rankings greatly, but they will improve the validity and acceptability of the reports.

Birth Certificates↗
PubMed1997

Influencing physician response to prenatal substance exposure through state legislation and work-place policies.

Little research attention has focused on ways to encourage physician response to prenatal substance exposure. We report initial results from a study examining the impact of state laws and work-place policies on physician response by combining legal analyses and data from a national physician survey. Our findings indicate that the message that laws and policies exist usually does not reach physicians. However, when the message does come through, some physician behaviors are influenced. In particular, physicians in states with clearer policies and behavioral expectations are significantly more likely to know and understand the law than physicians in other states. Further, believing that a work-place protocol on prenatal substance exposure exists is associated with significantly increased likelihood of an active response in case vignettes portraying prenatal substance exposure. The findings suggest that state legislative behaviors may increase physician response to prenatal substance exposure, but that response depends on the nature of the policy and on efforts to disseminate it.

California↗
PubMed1996

Protein kinase C-dependent regulation of human erythroleukemia (HEL) cell sphingosine kinase activity.

Sphingosine kinase functions in both the catabolism of sphingosine and in signal transduction pathways utilizing sphingosine-1-phosphate. The regulation of sphingosine kinase activity in human erythroleukemia (HEL) cells was investigated by treatment with several bioactive agents. Treatment of HEL cells with phorbol 12-myristate 13-acetate (PMA) caused a time- and concentration-dependent increase in sphingosine kinase activity measured in vitro. Sphingosine kinase activity increased in a phorbol ester- and diacylglycerol-specific manner. Staurosporine and calphostin C, protein kinase C (PKC) inhibitors, blocked the increased in sphingosine kinase activity, suggesting a PKC-dependent regulation. The effects of PMA on sphingosine kinase were dependent on transcription and translation. Purified PKC had no direct effect on sphingosine kinase activity. However, these studies led to the observation that HEL cell sphingosine kinase activity is stimulated in vitro by phosphatidylserine. Interestingly, other inducers of HEL cell differentiation, dimethylsulfoxide and retinoic acid, did not affect sphingosine kinase activity. These results indicate a separate and distinct pathway of PKC-dependent sphingosine kinase activation, and suggest a role for sphingosine kinase in regulation of cell function.

Cell Differentiation↗
PubMed1996

Effects of sphingosine stereoisomers on P-glycoprotein phosphorylation and vinblastine accumulation in multidrug-resistant MCF-7 cells.

To investigate the role of protein kinase C (PKC) in the regulation of multidrug resistance and P-glycoprotein (P-gp) phosphorylation, the natural isomer of sphingosine (SPH), D-erythro sphingosine (De SPH), and its three unnatural stereoisomers were synthesized. The SPH isomers showed similar potencies as inhibitors of in vitro PKC activity and phorbol binding, with IC50 values of approximately 50 microM in both assays. Treatment of multidrug-resistant MCF-7ADR cells with SPH stereoisomers increased vinblastine (VLB) accumulation up to 6-fold at 50 microM but did not alter VLB accumulation in drug-sensitive MCF-7 wild-type (WT) cells or accumulation of 5-fluorouracil in either cell line. Phorbol dibutyrate treatment of MCF-7ADR cells increased phosphorylation of P-gp, and this increase was inhibited by prior treatment with SPH stereoisomers. Treatment of MCF-7ADR cells with SPH stereoisomers decreased basal phosphorylation of the P-gp, suggesting inhibition of PKC-mediated phosphorylation of P-gp. Most drugs that are known to reverse multidrug resistance, including several PKC inhibitors, have been shown to directly interact with P-gp and inhibit drug binding. SPH stereoisomers did not inhibit specific binding of [3H] VLB to MCF-7ADR cell membranes or [3H]azidopine photoaffinity labeling of P-gp or alter P-gp ATPase activity. These results suggest that SPH isomers are not substrates of P-gp and suggest that modulation of VLB accumulation by SPH stereoisomers is associated with inhibition of PKC-mediated phosphorylation of P-gp.

ATP Binding Cassette Transporter, Subfamily B, Mem↗
PubMed1996

The solution structure of the Raf-1 cysteine-rich domain: a novel ras and phospholipid binding site.

The Raf-1 protein kinase is the best-characterized downstream effector of activated Ras. Interaction with Ras leads to Raf-1 activation and results in transduction of cell growth and differentiation signals. The details of Raf-1 activation are unclear, but our characterization of a second Ras-binding site in the cysteine-rich domain (CRD) and the involvement of both Ras-binding sites in effective Raf-1-mediated transformation provides insight into the molecular aspects and consequences of Ras-Raf interactions. The Raf-1 CRD is a member of an emerging family of domains, many of which are found within signal transducing proteins. Several contain binding sites for diacylglycerol (or phorbol esters) and phosphatidylserine and are believed to play a role in membrane translocation and enzyme activation. The CRD from Raf-1 does not bind diacylglycerol but interacts with Ras and phosphatidylserine. To investigate the ligand-binding specificities associated with CRDs, we have determined the solution structure of the Raf-1 CRD using heteronuclear multidimensional NMR. We show that there are differences between this structure and the structures of two related domains from protein kinase C (PKC). The differences are confined to regions of the CRDs involved in binding phorbol ester in the PKC domains. Since phosphatidylserine is a common ligand, we expect its binding site to be located in regions where the structures of the Raf-1 and PKC domains are similar. The structure of the Raf-1 CRD represents an example of this family of domains that does not bind diacylglycerol and provides a framework for investigating its interactions with other molecules.

Amino Acid Sequence↗
PubMed1996

Raf-1 kinase possesses distinct binding domains for phosphatidylserine and phosphatidic acid. Phosphatidic acid regulates the translocation of Raf-1 in 12-O-tetradecanoylphorbol-13-acetate-stimulated Madin-Darby canine kidney cells.

Previous studies demonstrated that the cysteine-rich amino-terminal domain of Raf-1 kinase interacts selectively with phosphatidylserine (Ghosh, S., Xie, W. Q., Quest, A. F. G., Mabrouk, G. M., Strum, J. C., and Bell, R. M. (1994) J. Biol. Chem. 269, 10000-10007). Further analysis showed that full-length Raf-1 bound to both phosphatidylserine and phosphatidic acid (PA). Specifically, a carboxyl-terminal domain of Raf-1 kinase (RafC; residues 295 648 of human Raf-1) interacted strongly with phosphatidic acid. The binding of RafC to PA displayed positive cooperativity with Hill numbers between 3.3 and 6.2; the apparent Kd ranged from 4.9 +/- 0.6 to 7.8 +/- 0.9 mol % PA. The interaction of RafC with PA displayed a pH dependence distinct from the interaction between the cysteine-rich domain of Raf-1 and PA. Also, the RafC-PA interaction was unaffected at high ionic strength. Of all the lipids tested, only PA and cardiolipin exhibited high affinity binding; other acidic lipids were either ineffective or weakly effective. By deletion mutagenesis, the PA binding site within RafC was narrowed down to a 35-amino acid segment between residues 389 and 423. RafC did not bind phosphatidyl alcohols; also, inhibition of PA formation in Madin-Darby canine kidney cells by treatment with 1% ethanol significantly reduced the translocation of Raf-1 from the cytosol to the membrane following stimulation with 12-O-tetradecanoylphorbol-13-acetate. These results suggest a potential role of the lipid second messenger, PA, in the regulation of translocation and subsequent activation of Raf-1 in vivo.

Amino Acid Sequence↗
PubMed1996

Peptides containing a consensus Ras binding sequence from Raf-1 and theGTPase activating protein NF1 inhibit Ras function.

A key event in Ras-mediated signal transduction and transformation involves Ras interaction with its downstream effector targets. Although substantial evidence has established that the Raf-1 serine/threonine kinase is a critical effector of Ras function, there is increasing evidence that Ras function is mediated through interaction with multiple effectors to trigger Raf-independent signaling pathways. In addition to the two Ras GTPase activating proteins (GAPs; p120- and NF1-GAP), other candidate effectors include activators of the Ras-related Ral proteins (RalGDS and RGL) and phosphatidylinositol 3-kinase. Interaction between Ras and its effectors requires an intact Ras effector domain and involves preferential recognition of active Ras-GTP. Surprisingly, these functionally diverse effectors lack significant sequence homology and no consensus Ras binding sequence has been described. We have now identified a consensus Ras binding sequence shared among a subset of Ras effectors. We have also shown that peptides containing this sequence from Raf-1 (RKTFLKLA) and NF1-GAP (RRFFLDIA) block NF1-GAP stimulation of Ras GTPase activity and Ras-mediated activation of mitogen-activated protein kinases. In summary, the identification of a consensus Ras-GTP binding sequence establishes a structural basis for the ability of diverse effector proteins to interact with Ras-GTP. Furthermore, our demonstration that peptides that contain Ras-GTP binding sequences can block Ras function provides a step toward the development of anti-Ras agents.

3T3 Cells↗
PubMed1996

Phospholipid and cation activation of chimaeric choline/ethanolamine phosphotransferases.

The Saccharomyces cerevisiae CPT1 and EPT1 genes encode for a cholinephosphotransferase (CPT) and choline/ethanolaminephosphotransferase, respectively. Both Cpt1p and Ept1p activities display an absolute requirement for cations and phospholipids. A mixed-micelle assay was employed to determine cation and lipid activators of parental and chimaeric Cpt1p/Ept1p enzymes to gain insight into their mechanism(s) of activation. Mg2+, Mn2+ and Co2+ were the only cations capable of activating Cpt1p and Ept1p in vitro. Kinetic data revealed that only Mg2+ is present in appropriate amounts to activate CPT activity in vivo. Kinetic data revealed that only Mg2+ is present in appropriate amounts to activate CPT activity in vivo. The two enzymes displayed distinct activation profiles on the basis of their relative affinities for Mg2+, and Mn2+ and Co2+. This allowed the use of chimaeric enzymes to determine the mechanism of cation activation. Cations do not activate Cpt1p or Ept1p by complexing with the substrate, CDP-choline, but instead bind to disparate regions within the enzymes themselves. Cpt1p and Ept1p also displayed distinct phospholipid activation profiles. Phospholipid activation required a phosphate and/or glycero-phosphoester linkage, with the phospho-head group moiety positioned at the surface of the micelle. Assays with parental and chimaeric Cpt1p/Ept1p constructs revealed that the phospholipid binding/activation domains are not located within linear segments of the protein, but instead are contained within distinct and separate regions of the proteins that require an intact tertiary structure for formation. Phosphatidylcholine (and its structural analogue sphingomyelin) were the best lipid activators of Cpt1p, the main biologically relevant CPT activity in S. cerevisiae. Hence CPT displays product activation. Because phosphatidylcholine is an efficient activator of CPT activity (and hence Cpt1p is not subject to feedback inhibition by its product), Cpt1p is incapable of functioning as a direct monitor of membrane phosphatidylcholine composition.

Cations, Divalent↗
PubMed1996

Communication between adolescents and physicians about sexual behavior and risk prevention.

OBJECTIVES: To assess the extent to which adolescents in a nonclinical community-based population have talked with a physician about sexual behavior and risk prevention and to examine whether adolescents value these discussions and trust physicians to protect their confidentiality. DESIGN: Self-administered anonymous survey. SETTING: Urban California school district. PARTICIPANTS: A total of 2026 students in 9th to 12th grade, 98% of the eligible students present on the survey day. OUTCOME MEASURES: Discussions with physicians about sexual matters, helpfulness of discussions, trust in physicians to protect confidentiality, and knowledge about confidentiality laws. RESULTS: Thirty-nine percent of adolescents reported discussions with physicians about how to avoid getting acquired immunodeficiency syndrome from sex, 37% about using condoms for vaginal intercourse, 13% about how to use condoms, 15% about the adolescent's sex life, 13% about how to say no to unwanted sex, and 8% about sexual orientation. In addition, 8% of adolescents had been given a condom by a physician. Adolescents were more likely to report most of these topics if they had ever had vaginal intercourse or if they had a regular physician. Most adolescents (80%-90%) would find it at least a little helpful to talk with a physician about various sexual matters. Most would trust a physician to keep secret that they asked questions about sex (75%), that they were having sex (65%), or that they were using contraception (68%). Fewer would trust physicians to keep secret a sexually transmitted disease (44%) or pregnancy (44%). For adolescents who knew that physicians in their state do not have to tell parents about sexually transmitted diseases or pregnancy, levels of trust rose, but only to 54%. CONCLUSIONS: Although professional medical organizations recommend that physicians discuss sexual matters and risk prevention with their adolescent patients, most adolescents report not having received these services. Physicians should be more aggressive about discussing these topics.

Adolescent↗
PubMed1996

Teenagers and alcohol misuse in the United States: by any definition, it's a big problem.

Despite the fact that more adolescents use alcohol than any other drug, studies of teenage alcohol misuse are relatively rare. Those that exist frequently fail to include high school dropouts and often focus on only part of the problem, such as how much or how often the adolescent drinks. This study examines the prevalence and demographic predictors of teenage alcohol misuse in a diverse sample of 4390 high school seniors and dropouts. It focuses on three different dimensions of misuse--high-risk drinking, alcohol-related problems and high consumption--and provides prevalence estimates by gender and race/ethnicity that are weighted to represent the original seventh grade cohort of 30 California and Oregon schools. Results show that by grade 12, nearly 70% of these teenagers have exhibited some form of alcohol misuse within the past year. Two-thirds have engaged in high-risk drinking and over 50% have experienced one or more alcohol-related problems. More stringent estimates that require variety or persistence of risky drinking and/or alcohol-related problems still capture between 40% and 54% of this population. However, focusing solely on high consumption fails to identify as many as half of these at-risk misusers. Males and females both exhibit high rates of alcohol misuse, as do most racial ethnic groups. However, African Americans and Asians are less likely to misuse alcohol than whites and Hispanics. The results underscore the need for including different forms of alcohol misuse in prevention programs, for improving our understanding of its etiology, and for providing upper and lower bound estimates of alcohol misuse in future research.

Adolescent↗
PubMed1996

Pelvic trauma imaging: a blinded comparison of computed tomography and roentgenograms.

To determine the sensitivity for detecting pelvic pathology and instability, roentgenograms and computed tomographic (CT) scans from 59 patients with pelvic injuries that had been admitted to a Level I trauma center were randomly reviewed by a orthopedic surgeon blinded to the study. Normal control roentgenograms and CT scans were included to decrease observer bias. The anteroposterior (AP) roentgenogram detected 66% of all pelvic injuries, 78% of those involving the anterior ring, and 53% of those involving the posterior pelvic ring. The trauma CT scan, 10-mm axial images of the abdomen and pelvis, detected 86% of all pelvic injuries, and 78% of anterior ring and 93% of posterior ring injuries. The sensitivity for detecting pelvic instability from one plain film AP pelvis roentgenogram taken in a trauma room setting was 74%. Inlet and outlet views were 75% sensitive. Trauma CT scans were 93% sensitive and high-definition pelvic CT scans (5-mm pelvic cuts) yielded 100% sensitivity. The mechanism of injury could be ascertained with 73% sensitivity by plain films and with 79% sensitivity by inlet and outlet views; trauma CT scans were 96% and high-definition pelvic CT scans were 100% sensitive. When combined, the AP pelvis roentgenograms and trauma CT scans identified 96% of the injured structures and were 100% sensitive in determining injury force patterns and instability. The data suggested that a good quality AP pelvis roentgenogram in conjunction with a complete trauma CT scan of the abdomen/pelvis should identify both the injury mechanism and pelvic instability with a high degree of sensitivity.

Adolescent↗
PubMed1996

The sexual practices of adolescent virgins: genital sexual activities of high school students who have never had vaginal intercourse.

OBJECTIVES: The purpose of this study was to determine whether high school-aged virgins engage in sexual practices that can transmit sexually transmitted diseases, including the human immunodeficiency virus (HIV). METHODS: Data were collected from an anonymous self-administered survey of 2026 urban students in 9th through 12th grades. RESULTS: Forty-seven percent of adolescents were virgins (42% of male adolescents and 53% of female adolescents). Of those who were virgins, 29% and 31% reported that, during the prior year, they had engaged in heterosexual masturbation of a partner and masturbation by a partner, respectively. The corresponding rates for heterosexual fellatio with ejaculation, cunnilingus, and anal intercourse were 9%, 10%, and 1%. Homosexual sexual activities were rare. Condom use for fellatio was also rare. Level of risk of virgins' sexual activities was associated with illicit substance use and other non-sexual risk behaviors, even after demographic variables had been controlled. CONCLUSIONS: Few high school-aged virgins engaged in anal intercourse, but many engaged in other genital sexual activities. Some of these activities can transmit disease, and all can indicate a need for counseling about sexual decision making, risk, and prevention.

Adolescent↗
PubMed1995

Regulation of phospholipid biosynthesis in Saccharomyces cerevisiae by CTP.

In the yeast Saccharomyces cerevisiae, the major membrane phospholipid phosphatidylcholine is synthesized by the CDP-diacylglycerol and CDP-choline pathways. We examined the regulation of phosphatidylcholine synthesis by CTP. The cellular concentration of CTP was elevated (2.4-fold) by overexpressing CTP synthetase, the enzyme responsible for the synthesis of CTP. The overexpression of CTP synthetase resulted in a 2-fold increase in the utilization of the CDP-choline pathway for phosphatidylcholine synthesis. The increase in CDP-choline pathway usage was not due to an increase in the expression of any of the enzymes in this pathway. CDP-choline, the product of the phosphocholine cytidylyltransferase reaction, was the limiting intermediate in the CDP-choline pathway. The apparent Km of CTP (1.4 mM) for phosphocholine cytidylyltransferase was 2-fold higher than the cellular concentration of CTP (0.7 mM) in control cells. This provided an explanation of why the overexpression of CTP synthetase caused an increase in the cellular concentration of CDP-choline. Phosphatidylserine synthase activity was reduced in cells overexpressing CTP synthetase. This was not due to a transcriptional repression mechanism. Instead, the decrease in phosphatidylserine synthase activity was due, at least in part, to a direct inhibition of activity by CTP. These results show that CTP plays a role in the regulation of the pathways by which phosphatidylcholine is synthesized. This regulation includes the supple of CTP for the phosphocholine cytidylyltransferase reaction in the CDP-choline pathway and the inhibition of the phosphatidylserine synthase reaction in the CDP-diacylglycerol pathway.

CDPdiacylglycerol-Serine O-Phosphatidyltransferase↗
PubMed1995

Ceramide triggers meiotic cell cycle progression in Xenopus oocytes. A potential mediator of progesterone-induced maturation.

The role of sphingomyelin-derived second messengers in progesterone-induced reinitiation of the meiotic cell cycle of Xenopus laevis oocytes was investigated. A brief treatment of defolliculated oocytes with sphingomyelinase (Staphylococcus aureus) was sufficient to induce maturation as measured by H1 kinase activity and germinal vesicle breakdown (GVBD). Pretreatment with cycloheximide inhibited sphingomyelinase-induced GVBD demonstrating a requirement for protein synthesis. Microinjection of ceramide or sphingosine, potential products of sphingomyelin hydrolysis, were capable of inducing GVBD in the absence of hormone. Metabolic labeling studies suggested the conversion of sphingosine to ceramide was necessary for sphingosine-induced GVBD. Additionally, fumonisin b1, an inhibitor of sphingosine N-acyltransferase, blocked sphingosine-induced GVBD demonstrating that ceramide is the more proximal biologically active metabolite. Treatment of oocytes with progesterone, the physiological inducer of oocyte maturation, resulted in a time- and concentration-dependent increase in the mass of ceramide and decrease in the mass of sphingomyelin through activation of a Mg(2+)-dependent neutral sphingomyelinase. These observations suggest that the generation of ceramide from sphingomyelin is part of the signal transduction pathway activated in response to progesterone and that the increase in ceramide is likely to be functionally important in resumption of the meiotic cell cycle.

Amino Acid Sequence↗