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Biomedical subjects

J Rivera

Publications and source records attributed to J Rivera.

At least 163 records · Page 9Linked to original sources
PubMed1995

Regulation of the adapter molecule Grb2 by the Fc epsilon R1 in the mast cell line RBL2H3.

Antigenic cross-linking of the high affinity IgE receptor (Fc epsilon R1) on mast cells results in protein tyrosine kinase activation. The object of the present study was to explore the regulation of the SH2 and SH3 domain containing adapter molecule Grb2 by Fc epsilon R1-stimulated PTK signal transduction pathways. Affinity purification of in vivo Grb2 complexes together with in vitro experiments with Grb2 glutathione S-transferase fusion proteins were used to analyze Grb2 complexes in the mast cell line RBL2H3. The data show that in RBL2H3 cells several different proteins are complexed to the SH3 domains of Grb2. These include the p21ras guanine nucleotide exchange factor Sos, two basally tyrosine-phosphorylated 110- and 120-kDa molecules, and a 75-kDa protein that is a substrate for Fc epsilon R1-activated PTKs. By analogy with Sos, p75, p110 and p120 are candidates for Grb2 effector proteins which suggests that Grb2 may be a pleiotropic adapter. Two Grb2 SH2-binding proteins were also characterized in RBL2H3 cells; the adapter Shc and a 33-kDa molecule. Shc is constitutively tyrosine phosphorylated in unstimulated cells and Fc epsilon R1 ligation induces no changes in its phosphorylation or binding to Grb2. In contrast, p33 is a substrate for Fc epsilon R1-activated PTKs and binds to Grb2 SH2 domains in Fc epsilon R1 activated but not quiescent cells. The beta subunit of the Fc epsilon R1 is a 33-kDa tyrosine phosphoprotein, but the p33 Grb2-binding protein described in the present report is not the Fc epsilon R1 beta chain and its identity is unknown. The present report thus demonstrates that there are multiple Grb2 containing protein complexes in mast cells of which a subset are Fc epsilon R1-regulated. Two other of the Grb2-binding proteins described herein are tyrosine phosphorylated in response to Fc epsilon R1 ligation: the 75-kDa protein which binds to Grb2 SH3 domains and the 33-kDa protein that associates with the Grb2 SH2 domain. We propose that protein complex formation by Grb2 is an important consequence of Fc epsilon R1 cross-linking and that this may be a signal transduction pathway which acts synergistically with calcium/PKC signals to bring about optimal mast cell end function.

Adaptor Proteins, Signal Transducing↗
PubMed1995

[The follow-up of the mobilization of circulating hematopoietic progenitor cells by granulocyte growth factor (G-CSF) using flow cytometry].

BACKGROUND: The aim of the present study was to investigate the characteristics of the mobilization of hematopoietic precursor cells CD34+ in peripheral blood following stimulation with recombinant granulocytic colony stimulating factor (G-CSF). METHODS: Fourteen patients (10 males, 4 females: mean age 33 years; range 14-58 years) diagnosed with oncohematologic neoplasms, in complete remission were studied. The patients had not received antineoplastic for at least four weeks prior to inclusion in the study. Recombinant G-CSF (8 micrograms/kg) was administered subcutaneously over a minimum of 4 days. Peripheral blood control were performed prior to administration of G-CSF (day 0), the third (day +3) day, and the sixth day (day +6). Daily leukapheresis was initiated at day +3 in 5 patients and at day +4 in 9 patients. The CD34+ cell content was determined in both peripheral blood and leukapheretic material by flow cytometry with an anti CD-34 monoclonal antibody conjugated with fluorescein. RESULTS: No significant differences were observed between the mononuclear cells and CD34+ counts obtained at the first apheresis and those performed at days +3 or +4 (32 +/- 14 x 10(9) vs 29 +/- 19 x 10(9) and 240 +/- 125 x 10(6) vs 162 +/- 160 x 10(6), respectively). The content of the apheresis products in CD34+ cells correlated positively with the number of these cells circulating in peripheral blood (r = 0.53, p = 0.001). In the second apheresis, the presence of mononuclear cells decreased approximately 20% with respect to the first, remained constant in later collections. To the contrary, a constant maintained decrease was observed in the collection of CD34+ on each leukaphesis in that the fourth apheresis only contributed in approximately 10% of the total quantity of CD34+ cells collected. CONCLUSIONS: Maximum mobilization of precursor cells was achieved on the third day at a dosage of 8 micrograms/kg/day, with the data found suggesting that three leukapheretic procedures are enough to collect most of the CD34+ cells mobilized.

Adolescent↗
PubMed1995

Effect of rhG-CSF on the mobilization of CD38 and HLA-DR subfractions of CD34+ peripheral blood progenitor cells.

Several studies have demonstrated that both CD34+/CD38- and CD34+/HLA-DR- human hematopoietic progenitor cells have properties associated with hematopoietic stem cells. However, the kinetics of these two cell populations in human peripheral blood (PB) after priming with granulocyte colony-stimulating factor (rhG-CSF) has not been investigated. By using flow-cytometric analysis we have shown that administration of rhG-CSF to 14 patients eligible for peripheral blood progenitor cell (PBPC) transplantation led to an increment of CD34+/CD38+ and CD34+/HLA-DR+ cells in the PB that paralleled the increase of total CD34+ cells, indicating that such subpopulations are responsible for the major release of CD34+ cells. Furthermore, rhG-CSF priming led to a significant mobilization of fractions of more immature CD34+/CD38- and CD34+/HLA-DR- cells to the PB. In the leukapheresis preparations, the average frequency of CD34+ cells lacking the CD38 or HLA-DR antigens was low (5% and 30%, respectively), with little overlap between the CD38- and HLA-DR- subpopulations. In addition, the yield of each subset of CD34+ cells (CD34+/CD38 +/- and CD34+/HLA-DR +/-) in the PB correlated with the numbers in the collected material. The results of the present study indicate that administration of rhG-CSF causes a significant increase of CD34+/CD38 +/- and CD34+/HLA-DR +/- cells in PB, and that such cells can be then safely harvested by leukapheresis procedures.

ADP-ribosyl Cyclase↗
PubMed1995

Misclassification of social disadvantage based on geographical areas: comparison of postcode and collector's district analyses.

BACKGROUND: Analysis of socioeconomic status (SES), defined on the basis of geographical area, will always be subject to misclassification of individuals. We studied the relationship between SES and selected health-related measures when SES was defined firstly on the basis of postcode and secondly on the basis of the smaller spatial area of collector's district (CD). METHOD: A Census population of 1.4 million was used to investigate the misclassification of individuals to SES group using postcode as opposed to CD. A field survey of 1000 respondents and a mortality register of 1756 deaths were used to compare the relationship between SES and certain outcome variables, when SES group was assigned using postcode and CD. Misclassification probability matrices were used to try to adjust the postcode-based results to approximate CD-based results. RESULTS: The Census data showed that nearly 50% of residents were misclassified into SES groups by the postcode-based system compared with a CD-based system. In comparing the most socially disadvantaged group with the least disadvantaged group, the postcode analysis underestimated the absolute effects of SES by 58% for the increased prevalence of smoking, by 19% for the reduced prevalence of participation in junior sporting clubs and by 13% for the increased mortality rate at ages 0-64 years. Adjustment of postcode-based results using misclassification matrices proved fruitless due to differential misclassification and technical difficulties. CONCLUSIONS: Misclassification of individuals to SES groups on the basis of postcode has caused an underestimation of the true relationship between SES and health-related measures. A reduction of this misclassification by using smaller spatial areas, such as CD or census enumeration districts, will provide improved validity in estimating the true relationship.

Adolescent↗
PubMed1995

Differential response to early nutrition supplementation: long-term effects on height at adolescence.

BACKGROUND: The classical risk approach to predicting who benefits from an intervention is unsound because it relies on the theoretical assumption that those at risk will necessarily benefit. A better approach to systematically test who benefits from nutrition supplementation is proposed using interactive models. METHODS: Differential effects of nutrition supplementation during early childhood on stature at adolescence were studied in 245 males and 215 females to identify determinants of long-term benefit from food supplementation. Factors studied included family socioeconomic status (SES) and children's home diet and diarrhoea during the first 3 years of life. To determine whether a factor conferred benefit, the statistical significance of the interaction between this factor and the intervention was tested. Data from the INCAP supplementation trial in Guatemala and from the follow-up of the same subjects at adolescence were used. RESULTS: Ordinary least squares (OLS) showed that high rates of diarrhoea in males and poor SES in females were significant determinants of benefit from supplementation at adolescence, and that the effects were mediated by length at 3 years old. Results of two-stage least squares (2SLS) analysis showed that length at 36 months, maturation and maternal height were significant determinants of height at adolescence but SES was not. CONCLUSIONS: Nutrition supplementation in early childhood has long-lasting effects on body size and the larger benefits acquired by some groups of children remain throughout early adulthood. The relevance of these findings for screening and targeting of nutritional interventions is discussed.

Adolescent↗
PubMed1995

Length screens better than weight in stunted populations.

Stunting (low length-for-age) is the most widespread manifestation of growth retardation worldwide. Yet, most nutrition programs use weight-for-age for screening of at-risk children. This study tested whether weight-for-age was an effective screening tool in a severely stunted rural Guatemalan population, using data from the INCAP longitudinal supplementation trial (n = 400). Stunting was defined as length-for-age < -2 SD of the National Center for Health Statistics standards at 3 y of age. Sensitivity and specificity analyses and receiver operating characteristics curves were used to compare weight indicators (weight-for-age, weight velocity and weight-for-length) with length (length-for-age and length velocity) and arm and head circumferences measured during early infancy. Length indicators were clearly superior to weight in predicting stunting (Zda test), and velocities were consistently worse than attained growth. Length-for-age at 6 mo had the best performance, followed by length-for-age at 3 mo, and weight-for-age at 6 and at 3 mo. Velocities, weight-for-length and circumferences were all poor predictors of stunting. Using the cutoff of < -1 SD, length-for-age at 3 mo was the best screening indicator for the early detection of growth faltering. Thus, the current use of weight-for-age, which results in large proportions of at-risk children being missed by screening, greatly limits the potential for impact of nutrition interventions.

Anthropometry↗
PubMed1995

Cryopreservation modifies flow-cytometric analysis of hemopoietic cells.

Although cryopreservation of human bone marrow has become very common in modern medicine, the knowledge on the effects of this procedure on hemopoietic cells is still limited. We herein have investigated whether the process of concentrating of human bone marrow to its buffy coat, the exposure to the cryoprotectant dimethyl sulfoide (DMSO), and/or the rate of controlled freezing/thawing procedures modifies the flow cytometric analysis of human bone marrow cells. We found that both the exposure of marrow cells to DMSO and/or the freezing procedure significantly modifies both the relative proportions of hemopoietic cell subsets and the intensity of expression of certain surface antigens. Thus, percentages of cells expressing CD7, CD13, CD33 and CD34, were found to be lower in both cryopreserved buffy coat and buffy coat merely exposed to DMSO, in comparison to those in untreated coat samples. Moreover, the intensity of the surface expression of CD33 decreased in cells exposed to DMSO, and further in cryopreserved samples. By contrast, the intensity of the CD19 antigen was higher in the last groups of samples than in the buffy coat or the unfractionated human bone marrow. Our present results suggest that flow-cytometric analysis of cryopreserved human bone marrow cells is not fully equivalent to that corresponding to fresh bone marrow or its fractionated buffy coat.

Antigens, CD↗
PubMed1995

Parturition: activation of stimulatory pathways or loss of uterine quiescence?

Parturition results from the establishment of phasic regular uterine contractions. Contractility in myometrial smooth muscle is stimulated by an increase in intracellular calcium ([Ca2+i]) which activates myosin light chain phosphorylation leading to increased myosin ATPase activity and enhanced rate of acto-myosin cross bridge formation. G proteins play a pivotal role in smooth muscle activation and relaxation by coupling cell membrane receptors to effector enzymes and ion channels. G alpha(s) and G alpha(i) stimulate and inhibit adenylyl cyclase, respectively and control cAMP formation. G alpha(q) stimulates phospholipase C resulting in the formation of two second messengers: inositol 1,4,5-trisphosphate (InsP3) which releases Ca2+ from the sarcoplasmic reticulum, and 1,2-diacylglycerol which activates protein kinase C. The oxytocin receptor stimulates myometrial contractility by increasing [Ca2+i] through both pertussis toxin resistant (G alpha(q)) and pertussis toxin sensitive (?G alpha (i)) pathways. beta-Adrenoceptors and prostaglandin EP2 receptors promote relaxation via G alpha(s)-adenylyl cyclase. The concentration of myometrial oxytocin receptors is five-times higher in pregnant compared to non-pregnant myometrium but decreases in samples obtained during labour. When myometrial slices are challenged with oxytocin there is a rapid increase in InsP3 levels with a time course which is similar to the rise in [Ca2+i] provoked by oxytocin in cultured myometrial cells. The formation of InsP3 in response to oxytocin in myometrial tissue at term is similar in samples obtained before and after the onset of labour. G alpha(q) and G alpha(i) are expressed at similar levels in non-pregnant and in pregnant myometrium obtained before or during labour. By contract, G alpha(s) levels are higher in pregnant compared to non-pregnant myometrium and decrease in samples obtained during labor. These changes in G alpha(s) are paralleled by prostaglandin E2-induced adenylyl cyclase activity in the same tissues. Parturition may be the consequence of downregulation of pathways that favour uterine quiescence by increasing cAMP formation, resulting in a relative dominance of stimulatory receptors that increase InsP3/Ca2+ availability.

Adenylyl Cyclases↗
PubMed1994

Phosphorylation of the gamma chain of the high affinity receptor for immunoglobulin E by receptor-associated protein kinase C-delta.

The gamma chain of the high affinity receptor for immunoglobulin E is a member of the T-cell antigen receptor zeta chain family and a functional subunit common to both T-cell and Fc receptors. Here we report that the gamma chain is phosphorylated on threonine in response to protein kinase C activation. Furthermore, the threonine phosphorylation of the gamma chain correlates with the endocytosis of this receptor. We identified a receptor-associated kinase as the calcium-independent protein kinase C-delta and found that it associates with the carboxyl-terminal cytoplasmic domain of the beta chain. In addition, protein kinase C-delta was the only isozyme capable of phosphorylating the gamma chain in vitro. These findings provide evidence for the functional role of protein kinase C-delta in early signal transduction events in the mast cell and suggest a more general mechanism of activation for receptors that share subunits of the zeta chain family.

Animals↗
PubMed1994

Protein kinases C-beta and C-epsilon link the mast cell high-affinity receptor for IgE to the expression of c-fos and c-jun.

In this report we identify the specific isozymes of protein kinase C (PKC) that are involved in c-fos and c-jun mRNA accumulation in the rat basophilic leukemia cell line RBL-2H3. These cells could be largely depleted of the endogenous PKC isozymes by chronic treatment with phorbol 12-myristate 13-acetate followed by permeabilization of the cells with streptolysin O. The reconstitution of these cells with defined concentrations of either PKC-beta or PKC-epsilon up to 10 nM and 20 nM, respectively, induced c-fos and c-jun in a dose-dependent manner. At high concentrations of PKC-beta and -epsilon the induction of c-fos and c-jun was independent of the aggregation of the high-affinity IgE receptors (Fc epsilon type I receptors). In contrast, at limiting concentrations of these two PKC isozymes, 1 nM, the increase in c-fos and c-jun mRNAs was dependent on the aggregation of the Fc epsilon type I receptors. Unlike PKC-beta and -epsilon, PKC-alpha and PKC-delta failed to reconstitute c-fos and c-jun induction at any dose over the range examined. We conclude that PKC-beta and PKC-epsilon serve as a link between the cell surface receptor and gene expression.

Animals↗
PubMed1994

Quantitative defect of glycoprotein Ib in severe cirrhotic patients.

A decrease of platelet agglutination induced by ristocetin has been described in cirrhotic patients. In order to investigate the relationship of such phenomenon with a putative defect on platelet membrane glycoprotein Ib, we have studied the quantitative and functional status of Gp Ib in eleven severe alcoholic cirrhotic patients, and the ability of their formalin-fixed platelets to agglutinate in the presence of normal plasma plus ristocetin. Interestingly, we found a significant decrease of immunoreactive GP Ib molecules (9,978 +/- 1,534 vs. 17,064 +/- 404 molecules per platelet) and ristocetin-dependent binding of vWF (9,113 +/- 1,338 vs. 13,992 +/- 1,968 molecules per platelet) (P < 0.01) in comparison to the levels found in a control group of healthy subjects. Immunoblotting analysis of platelet lysates confirmed the reduction of GP Ib level in cirrhotic patients, but showed no modification on the precipitation pattern of this glycoprotein. The ristocetin-induced platelet agglutination (light transmission %) was also significantly lower in patients with cirrhosis (48 +/- 7.6 vs. 92 +/- 3.6, P < 0.01), and correlated with the binding of normal vWF (r = 0.863, P = 0.0013). Patients with bleeding times longer than 7 min and/or clinical history of bleeding episodes showed the lowest values of platelet agglutination and GP Ib level. In conclusion, our present results indicate that liver cirrhosis is associated with a relevant decrease of functional GP Ib molecules on the platelet surface. This reduction might be related to the prolongation of bleeding time and to the bleeding diathesis observed in cirrhotic patients.

Adult↗
PubMed1994

Stability of glycoproteins Ib/IX and IIb/IIIa during preparation and storage of platelet concentrates: detection by binding assays with epitope-defined monoclonal antibodies and physiological ligands.

Preservation of the glycoprotein (GP) complexes Ib/IX and IIb/IIIa, because of their role as specific platelet receptors for adhesive proteins, is essential for the haemostatic efficacy of transfusions of platelet concentrates (PCs). We have combined binding assays with epitope-defined monoclonal antibodies, and with the physiological ligands von Willebrand factor and fibrinogen (Fo), to investigate the total expression and functional status of these receptors, in PCs stored for up to 9 days. Routine preparation and storage for up to 5 days have no apparent effect on the surface expression and the functional status of GPs Ib/IX and IIb/IIIa. However, after prolonged storage for up to 9 days we found a 50% decrease in the level of the total and functional GP Ib/IX content of platelets. This finding parallelled a significant rise in plasma glycocalicin, a proteolytic fragment of GP Ib. In addition, long-term storage promoted an impairment in the exposure of GP IIb/IIIa to Fo, and a 50% decrease in Fo binding capacity, without affecting the complex quantitatively. Finally, we also noticed a storage-induced increase in the platelet surface expression of GMP 140, and after 9 days of storage there was a rise in mean platelet volume, and a significant reduction in pH levels.

Agglutination Tests↗