[Cerebral mechanisms involved in the interaction between pain and emotion]. / Mécanismes cérébraux impliqués dans l'interaction entre la douleur et les émotions.

INTRODUCTION: Pain is an unpleasant and intrusive sensation, warning of actual or potential tissue damage. Over the last fifteen years, functional cerebral imaging research has demonstrated the involvement of many cerebral structures in the experience of pain. BACKGROUND: Intimately linked to the notion of suffering, the affective dimension of pain relies on neurophysiological systems partly distinct anatomically from those involved more specifically in its sensory dimension. Some pathways convey nociceptive information to the somatosensory cortex and the insula, contributing to the sensory aspects of pain (e.g.: sensory intensity), and secondarily, to its affective dimension. Other pathways project directly to the anterior cingulate cortex, the insula, the amygdala and to the prefrontal cortices, which are structures involved in the affective dimension of pain (unpleasantness of pain and regulation of autonomic and behavioral responses). Interestingly, these latter regions are an integral part of the cerebral emotional networks. PERSPECTIVES AND CONCLUSION: This close anatomical relationship between pain and emotions circuits could explain the powerful emotional impact of pain as well as the reciprocal modulatory effect of emotions on pain observed in clinical and experimental studies. More specifically, this modulatory effect might reflect interactions between emotional and nociceptive systems in the prefrontal and cingulate cortices, ventral striatum, amygdala and hippocampal regions. Taken together, these observations further attest to the emotional nature of pain experience.

Thrombospondin-1 gene expression affects survival and tumor spectrum of p53-deficient mice.

In vitro and in vivo data indicate that thrombospondin-1 (TSP1) inhibits tumor progression in several ways including direct effects on cellular growth and apoptosis in the stromal compartment. To evaluate the importance of TSP1 for the progression of naturally arising tumors in vivo, we have crossed TSP1-deficient mice with p53-deficient mice. In p53-null mice, the absence of TSP1 decreases survival from 160 +/- 52 days to 149 +/- 42 days. A log-rank test comparing survival curves for these two populations yields a two-sided P value of 0.0272. For mice that are heterozygous for the p53-null allele, survival is 500 +/- 103 days in the presence of TSP1 expression, and 426 +/- 125 days in its absence (P = 0.0058). Whereas TSP1 expression did not cause a measurable change in the incidence of the majority of tumor types, a statistically significant (P < or = 0.05) decrease in the incidence of osteosarcomas is observed in the absence of TSP1. To determine more directly if host TSP1 inhibits tumor growth, B16F10 melanoma and F9 testicular teratocarcinoma cells have been implanted in C57BL/6J and 129Sv TSP1-null mice, respectively. The B16F10 tumors grow approximately twice as fast in the TSP1-null background and exhibit an increase in vascular density, a decrease in the rate of tumor cell apoptosis, and an increase in the rate of tumor cell proliferation. Increased tumor growth is also observed in the absence of TSP1 on the 129Sv genetic background. These data indicate that endogenous host TSP1 functions as a modifier or landscaper gene to suppress tumor growth.

Thrombospondin-1 type 1 repeat recombinant proteins inhibit tumor growth through transforming growth factor-beta-dependent and -independent mechanisms.

Thrombospondin-1 (TSP-1) is a potent inhibitor of tumor growth and angiogenesis. The antiangiogenic activity of TSP-1 has been mapped to the procollagen homology region and the type 1 repeats (TSR) using synthetic peptides. To elucidate the molecular mechanisms that are involved in the inhibition of tumor growth by the TSRs, we have expressed recombinant versions of these motifs and have assayed their ability to inhibit the growth of experimental B16F10 melanomas and Lewis lung carcinomas. Recombinant proteins that contain all three TSRs (3TSR) or the second TSR with (TSR2+RFK) or without (TSR2) the transforming growth factor-beta (TGFbeta) activating sequence (RFK) have been expressed in Drosophila S2 cells. In addition, recombinant proteins with mutations in either the RFK sequence (TSR2+QFK) or the WSHWSPW sequence [TSR2 (W/T)] of the second TSR have been prepared. Similar to platelet TSP-1, these proteins are potent inhibitors of endothelial cell migration, and 3TSR of human TSP-1 (3TSR/hTSP-1) and TSR2+RFK activate TGFbeta. An 81% inhibition of B16F10 tumor growth is observed at 2.5 mg (135 nmol)/kg/day of the recombinant 3TSR/hTSP-1. A comparable level of inhibition is observed with 2.5 mg (360 nmol)/kg/day of TSR2+RFK. By contrast, 3TSR of mouse TSP-2 (3TSR/mTSP-2), TSR2+QFK, and TSR2 are significantly less effective. TSR2+RFK and TSR2 reduce tumor vessel density, but TSR2+RFK has a greater effect on B16F10 tumor cell apoptosis and proliferation. Concurrent treatment of B16F10 tumor-bearing mice with TSR2+RFK and either a soluble form of the TGFbeta receptor or an antibody to active TGFbeta reduces the inhibition of B16F10 tumor growth to levels that are comparable with those of TSR2 and TSR2+QFK. By contrast, the presence of the TGFbeta-activating sequence does not increase the level of inhibition of Lewis lung carcinoma experimental tumor growth. These data indicate that the TSRs inhibit tumor growth by inhibition of angiogenesis and regulation of tumor cell growth and apoptosis. The regulation of tumor cell growth and apoptosis is TGFbeta dependent, whereas the inhibition of angiogenesis is not.

Separation of root nodule cells and identification of tissue-specific genes.

The interior tissues of the alfalfa (Medicago sativa) root nodule differ in form and function from the peripheral layers. The interior tissues are specialized for the fixation of nitrogen in cells infected by rhizobia. In contrast, the peripheral nodule tissues perform roles that assist the interior tissues: they provide metabolic support and protect the interior tissues from damaging levels of oxygen. We used a novel microdissection technique to separate these tissue types, allowing immunological and molecular comparison between the nodule interior and periphery. Using differential mRNA display reverse transcription and polymerase chain reaction, we compared the mRNA profiles of the separated tissues and identified a transcript specific to the nodule interior, and several peripheral-specific candidate genes.

Thrombospondins in early Xenopus embryos: dynamic patterns of expression suggest diverse roles in nervous system, notochord, and muscle development.

The thrombospondins (TSPs) are a family of extracellular matrix (ECM) glycoproteins that modulate many cell behaviors including adhesion, migration, and proliferation. Here we report the molecular cloning of the Xenopus homologs of TSP-1 and TSP-3, and the developmental patterns of expression of Xenopus TSP-1, TSP-3, and TSP-4 mRNAs. Xenopus TSP-1 and TSP-3 protein sequences each share approximately 80% amino acid identity with their mammalian counterparts. TSP-1 mRNAs are detectable at low levels in fertilized eggs indicating that this TSP is a maternally deposited transcript. Zygotic expression of TSP-1, TSP-3, and TSP-4 begins at the end of gastrulation and transcripts encoding each protein accumulate through the tadpole stages of development. Whole mount in situ hybridizations reveal that each TSP mRNA is localized in the embryo with distinct, developmentally regulated patterns of expression. TSP-1 mRNAs are detected in a wide range of tissues including the floor plate of the neural tube, epidermis, somites, notochord and, most notably, alternating rhombomeres. Transcripts encoding TSP-3 are expressed in the notochord, floor plate, sensorial layer of the epidermis and sensory epithelia. TSP-4 mRNAs are restricted to somitic mesoderm and skeletal muscle. These data suggest that the TSPs represent a functionally diverse family of ECM proteins with tissue-specific functions during embryogenesis.

Thrombospondin-1 is required for normal murine pulmonary homeostasis and its absence causes pneumonia.

The thrombospondins are a family of extracellular calcium-binding proteins that modulate cellular phenotype. Thrombospondin-1 (TSP-1) reportedly regulates cellular attachment, proliferation, migration, and differentiation in vitro. To explore its function in vivo, we have disrupted the TSP-1 gene by homologous recombination in the mouse genome. Platelets from these mice are completely deficient in TSP-1 protein; however, thrombin-induced platelet aggregation is not diminished. TSP-1-deficient mice display a mild and variable lordotic curvature of the spine that is apparent from birth. These mice also display an increase in the number of circulating white blood cells, with monocytes and eosinophils having the largest percent increases. The brain, heart, kidney, spleen, stomach, intestines, aorta, and liver of TSP-1-deficient mice showed no major abnormalities. However, consistent with high levels of expression of TSP-1 in lung, we observe abnormalities in the lungs of mice that lack the protein. Although normal at birth, histopathological analysis of lungs from 4-wk-old TSP-1-deficient mice reveals extensive acute and organizing pneumonia, with neutrophils and macrophages. The macrophages stain for hemosiderin, indicating that diffuse alveolar hemorrhage is occurring. At later times, the number of neutrophils decreases and a striking increase in the number of hemosiderin-containing macrophages is observed associated with multiple-lineage epithelial hyperplasia and the deposition of collagen and elastin. A thickening and ruffling of the epithelium of the airways results from increasing cell proliferation in TSP-1-deficient mice. These results indicate that TSP-1 is involved in normal lung homeostasis.

Ability of the Higgins Nutrition Intervention Program to improve adolescent pregnancy outcome.

OBJECTIVES: To determine the extent to which birth weight can be increased and the risk for adverse pregnancy outcome decreased when pregnant adolescents participated in the Higgins Nutrition Intervention Program; and to describe the dietary components of the program, including their variation as a function of diagnosed risk for adverse pregnancy outcome. DESIGN: Retrospective cohort study involving review of medical charts. SUBJECTS/SETTING: Developed as an adjunct to routine prenatal care, the Higgins Nutrition Intervention Program consists of an assessment of each pregnant adolescent's risk profile for adverse pregnancy outcomes and an individualized nutritional rehabilitation program based on that profile. The intervention group for this evaluation consisted of 1,203 pregnant adolescents who participated in the Higgins program at the Montreal Diet Dispensary between 1981 and 1991. The nonintervention group consisted of a randomly selected group of 1,203 pregnant adolescents known not to have participated in the program. OUTCOMES MEASURED: Birth weight; rates of low birth weight, very low birth weight, preterm delivery, fetal growth retardation, perinatal morbidity and mortality; and maternal morbidity. STATISTICAL ANALYSIS: Means and proportions were used to describe risk profiles and pregnancy outcomes in the two groups. Analysis of covariance and logistic regression were used to compare pregnancy outcomes while controlling for the effect of key confounding variables. RESULTS: Results from multivariable analyses showed that infants in the intervention group weighed an average of 55 g more (P < .05) than infants in the nonintervention group; their low-birth-weight rate was 39% lower (P < .001) and their very-low-birth-weight rate was 56% lower (P < .01). Individually determined dietary prescriptions for the adolescents in the intervention group recommended increases in daily consumption averaging approximately 900 kcal energy and 52 g protein. The lowest daily increases (approximately 150 kcal energy and 2 g protein) were recommended to the group with no diagnosed risks; the greatest increases (approximately 1,300 kcal energy and 76 g protein) were recommended to the group with multiple risk conditions. Although none of the risk/intervention groups achieved their prescribed increases during intervention, increases in actual intake generally followed the pattern of the prescribed increases; that is, the greater the prescribed increase, the greater the actual increase. CONCLUSIONS: These results suggest that the Higgins Nutrition Intervention Program, in which nutrition intervention is individualized as a function of diagnosed risk, significantly improves the outcome of adolescent pregnancy.

Characterization of human thrombospondin-4.

The thrombospondins are a family of extracellular calcium binding proteins that are involved in cell proliferation, adhesion, and migration. We have sequenced full-length human thrombospondin-4 and characterized the recombinant protein. In contrast to Xenopus laevis thrombospondin-4, the human protein contains an RGD cell binding sequence in the third type 3 repeat. Transfection of mouse NIH3T3 fibroblasts or C2C12 myoblasts with a full-length human thrombospondin-4 cDNA results in the expression of a polypeptide with a reduced molecular weight of 140,000. In the absence of reducing agent, the expressed protein has an apparent molecular weight of 550,000. Recombinant thrombospondin-4 has been purified from the culture supernatant by heparin-Sepharose and anti-thrombospondin-4 antibody-Affi-gel affinity chromatography. Electron microscopy indicates that thrombospondin-4 is composed of five subunits with globular domains at each end. The observation of a calcium-dependent change in the electron microscopic appearance of thrombospondin-4 is consistent with limited tryptic digestion data that indicate that thrombospondin-4 is resistant to digestion in the presence of calcium. These data indicate that thrombospondin-4 is a pentameric protein that binds to heparin and calcium.

The evolution of the thrombospondin gene family.

Thrombospondin-1 is an adhesive glycoprotein that is involved in cellular attachment, spreading, migration, and proliferation. To date, four genes have been identified that encode for the members of the thrombospondin gene family. These four genes are homologous to each other in the EGF-like (type 2) repeats, the calcium-binding (type 3) motifs, and the COOH-terminal. The latter has been reported to be a cell-binding domain in thrombospondin-1. Phylogenetic trees have been constructed from the multisequence alignment of thrombospondin sequences from human, mouse, chicken, and frog. Two different algorithms generate comparable results in terms of the topology and the branch lengths. The analysis indicates that an early form of the thrombospondin gene duplicated about 925 million years ago. The gene duplication that produced the thrombospondin-1 and -2 branches of the family is predicted to have occurred 583 million years ago, whereas the gene duplication that produced the thrombospondin-3 and -4 branches of the family is predicted to have occurred 644 million years ago. These results indicate that the members of the thrombospondin gene family have existed throughout the evolution of the animal kingdom and thus probably participate in functions that are common to most of its members.

Identification and characterization of thrombospondin-4, a new member of the thrombospondin gene family.

A new member of the thrombospondin gene family, designated thrombospondin-4, has been identified in the Xenopus laevis genome. The predicted amino acid sequence indicates that the protein is similar to the other members of this gene family in the structure of the type 3 repeats and the COOH-terminal domain. Thrombospondin-4 contains four type 2 repeats and lacks the type 1 repeats that are found in thrombospondin-1 and 2. The amino-terminal domain of thrombospondin-4 has no significant homology with the other members of the thrombospondin gene family or with other proteins in the database. RNAse protection analysis establishes that the initial expression of Xenopus thrombospondin-4 is observed during neurulation. Levels of mRNA expression increase twofold during tailbud stages but decrease by the feeding tadpole stage. The size of the thrombospondin-4 message is 3.3 Kb and 3.4 Kb in the frog and human, respectively. Northern blot analysis of human tissues reveals high levels of thrombospondin-4 expression in heart and skeletal muscle, low levels in brain, lung and pancreas and undetectable levels in the placenta, liver and kidney. These data establish the existence of a new member of the thrombospondin gene family that may participate in the genesis and function of cardiac and skeletal muscle.

Expression and mutagenesis of thrombospondin.

Thrombospondin is a 420,000-dalton adhesive glycoprotein that is composed of three subunits of equivalent molecular weight. When the cDNA for the complete coding region of the human endothelial cell thrombospondin subunit is expressed in mouse NIH 3T3 cells, a 420,000-dalton protein is synthesized and secreted. The expressed protein comigrates with human platelet thrombospondin both in the presence and in the absence of a reducing agent. The expressed protein binds to a monoclonal anti-thrombospondin antibody, heparin, and calcium. In addition to the 420,000-dalton protein, the transfected cell lines also express a variable amount of a 140,000-dalton polypeptide. When the culture supernatants that are produced by cells that are expressing thrombospondin are applied to heparin-Sepharose, the 420,000-dalton and the 140,000-dalton proteins are bound to the column and are eluted with buffer containing 0.55 and 0.3 M NaCl, respectively. The 140,000-dalton protein only binds to heparin-Sepharose in the presence of calcium. Deletion of the region of homology with procollagen results in defective assembly of the trimer. Deletion of the type 1 or type 2 repeats results in decreased stability of the subunit with the predominant polypeptides that are expressed having molecular weights of 127,000 and 130,000, respectively. These polypeptides retain low-affinity heparin-binding activity. High-affinity heparin binding is markedly diminished by mutations in either of two sequence motifs that include clusters of lysines and arginines.(ABSTRACT TRUNCATED AT 250 WORDS)

Characterization of the murine thrombospondin gene.

Thrombospondin is an adhesive glycoprotein that supports cell attachment, spreading, and migration. The murine thrombospondin gene is approximately 18 kb in length and includes 22 exons. Interspecific backcross analysis using progeny derived from matings of (C57BL/6J x Mus spretus) F1 x C57BL/6J mice indicates that the thrombospondin gene is tightly linked to the Fshb, Actcl, Ltk, and B2M loci on murine chromosome 2. The sequence of the murine gene is very similar to that of the human gene in (1) regions of the promoter, (2) the coding region, and (3) the 3'-untranslated region. The predicted amino acid sequence of the mature murine thrombospondin subunit is 95.1% identical to that of the human. The sequences of these two species are most similar at the regions containing the type 1, 2, and 3 repeats as well as the COOH-terminal globular domain. The thrombospondin promoter is similar to the 5' flanking region of some housekeeping and growth control genes in that it contains multiple GC-rich regions and lacks a CAAT box. The presence of various consensus sequences suggests that thrombospondin gene expression is regulated by cAMP, cytokines, and steroid hormones.

Twin pregnancy: the impact of the Higgins Nutrition Intervention Program on maternal and neonatal outcomes.

Perinatal outcomes were compared between 354 twins treated with the Higgins Nutrition Intervention Program and 686 untreated twins. After differing distributions of key confounding variables were adjusted for, the twins in the intervention group weighed an average of 80 g more (P less than 0.06) than the nonintervention twins; their low-birth-weight rate was 25% lower (P less than 0.05) and their very-low-birth-weight rate was almost 50% lower (P less than 0.05). Although the rate of preterm delivery was 30% lower in the intervention group (P less than 0.05), the rates of intrauterine growth retardation were similar in the two groups. Fetal mortality was slightly higher (14 vs 12 per 1000, NS), but early neonatal mortality was fivefold lower (3 vs 19 per 1000, P less than 0.06) in the intervention group. Maternal morbidity was significantly lower (P less than 0.05) in the intervention group. There was a trend towards lower infant morbidity in the intervention group. These results suggest that nutritional intervention can significantly improve twin-pregnancy outcome.

Cloning and sequencing of chicken thrombospondin.

Thrombospondin is a multifunction adhesive protein with the ability to bind proteoglycans, cell surface receptors, other proteins, and calcium ions. Several sequence motifs for some of these interactions have been identified in human thrombospondin. To evaluate the potential functional significance of these sequences and to begin a study of the evolution of thrombospondin, we have isolated and sequenced thrombospondin cDNA clones from a chicken embryo library. Comparison of the chicken and human sequences reveals that the NH2-terminal heparin-binding domains are only 34% identical. By contrast, the type 3 repeats and the COOH-terminal domains are 80 and 82% identical, respectively, when comparing human and chicken sequences. Potential cell recognition sequences of RGD and VTCG are conserved, with the chicken sequence containing an additional copy of the VTCG sequence. Whereas substitutions occur in the two potential heparin-binding motifs that have human counterparts, the chicken sequence contains a third potential heparin-binding motif. The results indicate that the evolutionary constraints on the various types of cell-binding motifs may be quite different.

Detoxification and mineral supplementation as functions of geophagy.

Clays employed historically in the consumption of astringent acorns plus seven edible clays from Africa were examined in relation to the functional significance of human geophagy. On the basis of sorptive maxima for tannic acid ranging from 5.6 to 23.7 mg/g, we conclude that adsorption of tannic acid in traditional acorn preparation methods in California and Sardinia helped make these nuts palatable. Calcium available in solution at pH 2.0 and 0.1 mol NaCl/L was 2.10 and 0.71 mg/g for the Sardinian and Californian clays, respectively. The African clays released calcium, copper, iron, magnesium, manganese, or zinc in amounts of nutritional significance from some clays but not from others. A clay recovered from an archaeological site occupied by Homo erectus and early H. sapiens was indistinguishable mineralogically, in detoxification capacity and in available minerals, from clays used in Africa today. We suggest that the physiological significance of geophagy made it important in the evolution of human dietary behavior.

Separation of the lipid bilayer from the membrane skeleton during discocyte-echinocyte transformation of human erythrocyte ghosts.

The membrane skeleton, a protein lattice at the internal side of the red cell membrane, is principally composed of spectrin, actin and proteins 4.1 and 4.9. We have examined negatively stained red cell ghosts and demonstrated, on an ultrastructural level, a separation of the lipid bilayer from the membrane skeleton during echinocytic transformation. The electron micrographs of discoidal red cell ghosts suspended in hypotonic buffer revealed a filamentous reticulum that uniformly laminated the entire submembrane region. transformation of the discoidal ghosts into echinocytic form, as induced by incubation in isotonic buffer, resulted in a disruption of skeletal continuity underlying the surface contour of the membrane spicule. The submembrane reticulum extended into the base and the neck of the spiny processes of the crenated ghosts but was absent at the tip of these projections. In addition, membrane vesicles without a submembrane reticulum were detected either attached to the tips of the spicules or released into the supernatant from the echinocytic ghosts. Protein analysis revealed that the released vesicles were enriched in bands 3, 4.1 and 7 and contained very little of the membrane skeletal proteins, spectrin and actin. The data indicate that during echinocyte formation, parts of the lipid bilayer physically separate from the membrane skeleton, leading to a formation of skeleton-poor lipid vesicles.