Biomedical subjects

J Martinez

Publications and source records attributed to J Martinez.

At least 181 records · Page 10Linked to original sources

PubMed1998

Neurocircuitry of conditioned inhibition of analgesia: effects of amygdala, dorsal raphe, ventral medullary, and spinal cord lesions on antianalgesia in the rat.

Pain inhibition (analgesia) is produced by learned danger signals and inhibited by learned safety signals (antianalgesia). Conditioned analgesia is mediated by brain-to-spinal pathways releasing spinal endogenous opiates. Spinal morphine mimics learned danger signals in producing analgesia, which is inhibited by antianalgesia. The circuitry mediating antianalgesia is unknown. These experiments demonstrate that raphe dorsalis, raphe magnus, and spinal dorsolateral funiculus lesions abolish antianalgesia. Other lesions had no effect on antianalgesia. More important, lesions that blocked development of conditioned analgesia did not block development of antianalgesia. Thus, neural circuitries mediating analgesia and antianalgesia were found to be distinct, and conditioned inhibition of analgesia was found to act by inhibiting the most distal part of the conditioned analgesia circuit, namely, the spinal cord.

Amygdala↗

PubMed1998

Tenidap decreases IL-8 and monocyte chemotactic peptide-1 (MCP-1) mRNA expression in the synovial tissue of rabbits with antigen arthritis and in cultured synovial cells.

Since IL-8 and MCP-1 are chemoattractant proteins that participate in the recruitment of inflammatory cells into the arthritic joint, we examined the effects of tenidap, a new anti-inflammatory drug of the oxindole family, on IL-8 and MCP-1 expression in the joints of rabbits with acute antigen arthritis. The model was induced by injecting 5 mg/ml ovalbumin into the knees of 20 preimmunized rabbits. Animals were randomized into two groups: treated with tenidap (15 mg/kg per 12 h), or untreated. The effect of tenidap treatment was evaluated on chemokine production in synovial membranes of rabbits with arthritis and in cultured monocytic and synovial cells (SC). By immunoperoxidase staining, chemokines were localized in the synovial tissue. Chemokine messenger RNA levels in the synovial membranes and in cultured cells were analysed by reverse transcription-polymerase chain reaction (RT-PCR). At the end of the study, tenidap significantly reduced neutrophil infiltration into the joint cavity (27+/-4 x 10(6) cells/ml versus 45+/-6 x 10(6) cells/ml in untreated; P<0.05), and synovial effusion (134+/-15 microl versus 236+/-19 microl in untreated; P<0.005). Untreated rabbits showed synovial membrane up-regulation in mRNA expression of IL-8 and MCP-1 (11- and seven-fold versus healthy rabbits, respectively) that was markedly decreased by tenidap (two- and three-fold versus healthy rabbits, respectively). IL-8 and MCP-1 were localized in the synovial tissue in a perivascular pattern and areas of the interstitium and lining, mostly coinciding with cell infiltration. Tenidap also reduced the accumulation of IL-8 and MCP-1 proteins. In cultured synovial and monocytic cells, tumour necrosis factor-alpha (TNF-alpha) elicited an increase in gene expression of IL-8 (four- and nine-fold, respectively) and MCP-1 (nine- and four-fold, respectively) that was significantly reversed in both cell types by 10 microM tenidap. These results suggest that the beneficial effect of tenidap in acute antigen arthritis could be related to the down-regulation in gene expression and synthesis of IL-8 and MCP-1, two key chemokines involved in the recruitment of inflammatory cells.

Acute Disease↗

PubMed1998

Clinical experience in hospice and palliative medicine for clinicians in practice.

Many healthcare professionals already in practice have identified their need to pursue further practical training in the provision of hospice and palliative care. We began offering a 1-week clinical experience to physicians, nurses, pharmacists, social workers, and chaplains in the summer of 1995. As of October 1,1997, there have been 190 requests for application materials from individuals in more than 22 states, as well as from Singapore and Uganda. Thirty-five individuals completed visits by October 31,1997; 17 nurses, 16 physicians, 1 psychologist, and 1 chaplain. Although all are working in areas related to palliative care, 57% (20 of 35) were not currently working for a hospice program. A 25-question examination was administered as a needs-assessment test. Overall they scored 75% correct. They did especially poorly on questions related to dosing of opioids, assessment of pain, and prognosis in AIDS. They completed a videotaped interview with a standardized patient focusing on skills in discussing a terminal prognosis, "do not resuscitate" (DNR) status, and hospice referral. They evaluated the entire educational experience with a self-report at the end of their visit using a Likert Scale with values of 1 to 5. To the statement "I achieved the specific goals which I set for myself," the average score was 4.6 (range 1-5). To the statement "The experience was worth the time and effort," the average score was 4.9. To the statement "I would recommend this experience to others," the average score was 4.9. The evaluation was repeated 6 months after the visit with similar scores. In addition, to the statement "My current efforts are helping to change the way dying patients and their families are cared for in the broader environment in which I work," the average score was 4.9 (range 4-5). We conclude that this is a successful program of clinical exposure to hospice and palliative medicine for clinicians in practice.

Journal Article↗

PubMed1998

A program of hospice and palliative care in a private, nonprofit U.S. Teaching hospital.

A hospice and palliative care program was established at Northwestern Memorial Hospital, a 773-bed private nonprofit hospital located in the urban center of Chicago. The program consists of three components: consultation service, acute inpatient unit, and home-hospice program. The consultation service saw an average of 57 new patients per month (range 45-80) in fiscal year 1997. The 12-bed acute care inpatient unit had an average midnight census of 9.8 in fiscal 1996. This decreased to 6.9 in fiscal 1997 due to new treatment approaches for AIDS and has rebounded to nine in the third quarter of fiscal 1998. The unit cares for more than one third of dying patients in the hospital. Patients do not have to access hospice insurance benefits to be admitted to the unit. The home-hospice program has a median length of stay of 31 days and serves patients living within the city limits of Chicago. A total of 800 patients were referred to the program, and 370 patients died in the program in fiscal 1997. A total of 219 different physicians were attending physicians for patients in the program during a 3- year period. Revenue exceeded direct expenses by $1.48 million. Fee-for-service billing for physician services outside of those provided or billed under the Medicare Hospice Benefit are not included in these figures. The group practice that bills for the physicians collected an average of 50.5% of billed charges over 4 years. We conclude that a program of hospice and palliative care can be successful in a private teaching hospital in the United States.

Journal Article↗

PubMed1998

Prospective evaluation of referrals to a hospice/palliative medicine consultation service.

A hospice/palliative medicine consultation team was formed in July 1993 in a U.S. teaching hospital to provide patient care and education. The team consists of an attending physician, nurse, fellow, and any residents or medical students rotating on the service. More than 500 consultations are received each year. Beginning in January 1995,108 consecutive referrals to the service were assessed using a standard form completed by the nurse. The average age was 62 years. The gender of patients was 58% male and 42% female. At the time of consultation 87% were hospitalized on general medical services (including hematology/oncology), 4% were on surgical services, 3% on the neurology service, and 6% were in an intensive care unit. Cancer was the primary diagnosis in 52%, AIDS in 24%, with the rest being distributed among cardiac, renal, pulmonary, neurologic, and other diseases. The most prominent physical symptoms were 48% weakness/malaise, 44% pain, 28% dyspnea, and 23% agitation/confusion. The average length of time patients were followed was 2 days (range 1-10). We conclude that a hospice/palliative medicine consultation service sees a broad range of patients and problems and is a rich resource for teaching hospice and palliative medicine.

Journal Article↗

PubMed1998

Biological activity and three-dimensional structure of an agonist analog of bombesin.

JMV635, a nonapeptide analog of the active terminal nonapeptide segment of bombesin, was tested for its ability to stimulate in vitro amylase release from rat pancreatic acinar cells and to inhibit the binding of gastrin-releasing peptide to rat pancreatic acini. It was found to be a full agonist of bombesin and to recognize the bombesin receptor with moderate potency. The NMR proton assignments of JMV635 were achieved, and the conformations of JMV635 in aqueous solution and in trifluoroethanol at 297 K were determined using two-dimensional COSY, HOHAHA, NOESY and ROESY experiments. In trifluoroethanol, JMV635, like the active part of bombesin, showed a partial alpha-helical structure. These results were confirmed by circular dichroism and refined by restrained molecular dynamic methods. Structure calculations, using the distance and angle restraints obtained from NMR data on JMV635, gave a total of 75 structures which could be aligned to a root mean square deviation of the bond length of 0.007 A and of the valence angle of 1.55 degrees for the backbone atoms of the amino acid residues. The conformation is a well-defined right-handed alpha-helix in the C-terminal Q2-G6 segment and is less structured in the three C-terminal residues.

Amylases↗

PubMed1998

Synthesis and pharmacological evaluation of dimer derivatives of the bradykinin receptor antagonist HOE-140.

The synthesis and pharmacological evaluation of dimer derivatives of the C-terminal fragments of the potent bradykinin antagonist HOE-140, linked through their N-termini, were performed. The influence of peptide moiety length was studied using the succinyl moiety as a linker. Our attention focused on the dimer of the C-terminal tetrapeptide of HOE-140 (compound JMV 980), which displayed some inhibiting activity (IC50 = 247 nM) for bradykinin B2 receptors. Unexpectedly, it was orally active in inhibiting bradykinin-induced hypotension in the rat. Based on this tetrapeptide dimer model, we synthesized pseudotetrapeptide dimer bradykinin antagonists 29 and 33, which exhibited high affinity (Ki = 76 and 61 nM, respectively) for the human cloned B2 receptor. In addition, compound 29 inhibited bradykinin-induced contraction of the human umbilical vein giving a pKB value of 6.45. Compounds 29 and 33 were selective toward B2 receptors because they did not bind to the cloned human B1 receptor up to 10 microM.

Animals↗

PubMed1998

Cytokine responses in Trypanosoma cruzi-infected children in Paraguay.

Both parasite and host immune factors may contribute to the development and progression of chronic chagasic cardiomyopathy during Trypanosoma cruzi infections. The present study targeted infected children (5-14 years of age) from an endemic area of Paraguay in an analysis of T. cruzi-specific cytokine profiles. This age group is characteristically the most affected by the early phases of infection. Trypanosoma cruzi-induced cytokine gene expression (interleukin-2 [IL-2], and interferon-gamma [IFN-gamma], IL-4, and IL-10) was studied in 25 seropositive children categorized as being either acute, symptomatic, with Romana's sign (n = 2), or early, indeterminate (postacute, n = 23). Acutely infected children showed a distinct T helper cell-1 (Th1)-type (IFN-gamma) cytokine response to infection. The cytokine pattern that was observed in the seropositive, asymptomatic (early, indeterminate) group was of the Th0 type (expression of both IFN-gamma and IL-4). We hypothesize that selective induction of a Th0-type cytokine pattern is important for development of cell-mediated and humoral immune responses that suppress parasite burden, thereby prolonging the onset or limiting the severity of chronic Chagas' disease later in life.

Adolescent↗

PubMed1998

Youth counseled for HIV testing at school- and hospital-based clinics.

The prevalence of human immunodeficiency virus (HIV) in adolescents is difficult to assess as few adolescents consent to testing. This prospective study characterized urban youth requesting HIV testing at two types of health settings, inner-city school-based and hospital-based clinics. Data were obtained on 1652 inner-city youths aged 13 to 19 years who consented to individualized HIV counseling and testing from January 1993 to January 1994. Identified risks for HIV included sexual activity, sexually transmitted disease (STD) history, and substance use by self-report during a confidential structured interview. Data were analyzed using chi-squared analysis. Of the 1652 youth who were counseled, 1602 were from hospital-based clinics. A total of 827 (50%) requested HIV testing. Females accounted for the majority of youth who underwent counseling (79%) and requested HIV testing (75%). However, once counseled, males were more likely to be tested. Risk factors differed by gender; females were more likely to report STDs and marijuana use, and males more likely to report alcohol and cocaine use. These results indicate a need to identify developmentally appropriate methods to educate and counsel youth about HIV that will lead to more youth willing to be tested. School-based clinics may provide easier access than traditional health models for confidential HIV services.

AIDS Serodiagnosis↗

PubMed1998

Maturational changes in the thermal nociceptive responses of developing rats.

Some find developmental differences in rodent thermal nociceptive responses and others do not. To address these inconsistencies, the escape latencies of immature (5-to 25-day-old) and adult (3-to 4-month-old) albino rats were recorded following tail exposure to different intensities of radiant heat (650-W halogen lamp placed 10-30 mm from the tail) or conductive heat 35-50 degrees C water). Developmental differences in tail flick latencies were not observed in immature rats when the lamp was closest to the tail (although adult latencies were longer than 5-and 15-day-old responses) When radiant heat intensity was reduced, 5-day-old rats had shorter escape latencies than 15-, 25-, and 90-day-old animals. Age differences persisted in the latencies of immature animals even when the test aperture was varied to compensate for maturational changes in tail width (whereas adult responses no longer differed from those of 5-and 15-day-old rats). Developmental differences were eliminated when the tail skin was blackened so as to normalize the absorption of radiant heat across age. Similar age-and intensity-dependent differences were observed in rats exposed to conductive heat: Five-and 10-day-old pups had shorter escape responses than older rats when tails were immersed in intermediate (40 or 45 degrees C) but not lower (35 degree C) or higher (50 degrees C) temperature water. Blackening the tails did not change conductive heat escape latencies. No sex differences were found at any age or stimulus intensity with either type of heat. Higher intensities of thermal stimuli applied to the tail are required to elicit escape responses in older rats compared to younger ones, but the use of relatively intense thermal test stimuli can mask age-dependent differences in nociception. Some of the inconsistent results reported previously about maturational changes in thermal nociception may be due to intensity differences in the noxious test stimuli used. Maturational differences in the radiant absorption properties of the tail seem to account for most of the age-related changes in rodent responses to radiant heat, but the mechanism(s) which subserve developmental differences in conductive heat nociception need to be elucidated.

Age Factors↗

PubMed1998

Prevalence of micro- and macroalbuminuria in a Canarian population of type 2 diabetic patients. Relationship with blood pressure, lipid profile, obesity and metabolic control.

We performed a cross-sectional study on the prevalence of micro- and macroalbuminuria in a population of 288 Type 2 diabetic patients from Northern Gran Canaria Island (age 59 +/- 9.5, years; 48% male): 179 unselected patients referred by their family physicians, and 109 from our diabetes clinic. Sex, age, duration of diabetes and hypertension, blood pressure, body mass index, waist-hip ratio, HbA1c, creatinine, cholesterol (total and HDL), triglycerides, lipoprotein (a), and the presence of retinopathy, polyneuropathy, and coronary and cerebrovascular disease were assessed. The prevalences of micro- and macroalbuminuria were 28.5% and 11.8%. Among the patients referred by their family physicians, 32.4% were micro- and 6.1% macroalbuminuric. In our diabetes clinic, there were respectively 22% and 21% (with a higher prevalence of macroalbuminuria than in primary care, p < 0.05). Seventy-three percent were hypertensive in both settings. Prevalence was 31.5% for diabetic retinopathy, 21.0% for diabetic polyneuropathy, 8.1% for cerebrovascular disease, and 20.2% for coronary heart disease. The albumin excretion rate was significantly correlated with plasma creatinine levels, diastolic blood pressure, total cholesterol and the presence of coronary heart disease and diabetic retinopathy, but not with age, duration of diabetes or hypertension, body mass index, waist/hip ratio, glycated haemoglobin or triglycerides.

Albuminuria↗

PubMed1997

Melanin-concentrating hormone binding sites in human SVK14 keratinocytes.

Melanin concentrating hormone (MCH) is a cyclic peptide which regulates a broad array of functions in the mammalian brain and it may act as a paracrine factor in peripheral organs. In these studies a radiolabeled MCH derivative, the [125I]-[Phe13, Tyr19]-MCH, was synthesized and used as a tracer to perform binding experiments. A number of human or rodent cell lines displayed specific binding with [125I]-[Phe13, Tyr19]-MCH, the highest binding capacity being observed with human SVK14 keratinocytes. Saturation binding analysis with SVK14 cells indicated about 10,000 MCH binding sites per cell and a Kd of 0.7 nM for [125I]-[Phe13, Tyr19]-MCH. Surprisingly, the iodinated [Phe13, Tyr19]-MCH displayed about 10-fold higher affinity (Ki approximately 3.0 nM) for the putative MCH receptor than the noniodinated form (Ki approximately 25-30 nM). Competition binding analyses comparing various MCH-related peptides revealed a similar low binding potency for all these peptides (Ki approximately 65-160 nM). Strikingly, rat ANP and rat/human CNP but not rat BNP displaced [125I]-[Phe13, Tyr15]-MCH with Ki approximately 210-365 nM and may be due to topological similarities instead of partial sequence identities between MCH and some of the natriuretic peptides. However, other peptides such as CRF, alpha MSH, Arg-vasopressin, and MGOP-peptide I did not compete with the radioligand. Finally, the molecular mass of the MCH binding sites on SVK14 cells was estimated to be 47 kDa by crosslinking and SDS-PAGE experiments. Taken together, our data revealed the widespread expression of MCH binding sites on mammalian cells, particularly on skin carcinoma cells. However, the low affinity of these sites for the native MCH and MCH-related peptides as well as competitivity with ANP and CNP indicates that further biochemical and functional characterizations are needed to validate them as genuine physiological MCH receptors.

Amino Acid Sequence↗

PubMed1997

Effect of antipanic treatment on response to carbon dioxide.

BACKGROUND: Disordered breathing among patients with panic disorder, including hyperventilation during attacks and increased anxiogenic response to carbon dioxide (CO2) inhalation, is well established. We wished to assess whether there is a change in the physiological response to CO2 after patients have undergone antipanic therapy with either tricyclic antidepressants or cognitive behavioral therapy (CBT). METHODS: Twenty-nine patients with panic disorder underwent baseline CO2 sensitivity testing using the traditional Read rebreathing method and then received either antidepressant treatment (n = 21) or CBT (n = 8). After completing treatment, CO2 testing was repeated. A comparison sample of 14 normal volunteers also had two CO2 sensitivity tests, separated by an average of 21.6 (SD = 8.8) weeks. RESULTS: Using a liberal standard, in which all CO2 sensitivity tests whose correlations between minute ventilation and end-tidal CO2 were at least .75 were used, patients, but not controls, demonstrated a significant reduction in CO2 sensitivity between the first and second test. Using a more conservative .90 correlation standard reduced the sample size available and resulted in trend reduction in patients but no significant change in controls. There was a suggestion that the change was most pronounced in treatment responders, although the number of patient nonresponders is extremely small in this sample. CONCLUSIONS: These data indicate that treatment reduces CO2 sensitivity in patients with panic disorder. We speculate that manipulation of the serotonergic and noradrenergic neurotransmission systems, both known to play a role in the control of respiration, may have a specific effect in reducing respiratory hyperactivity in panic disorder.

Administration, Inhalation↗

PubMed1997

Differential targeting of nicotinic acetylcholine receptors by novel alphaA-conotoxins.

We describe the isolation and characterization of two peptide toxins from Conus ermineus venom targeted to nicotinic acetylcholine receptors (nAChRs). The peptide structures have been confirmed by mass spectrometry and chemical synthesis. In contrast to the 12-18 residue, 4 Cys-containing alpha-conotoxins, the new toxins have 30 residues and 6 Cys residues. The toxins, named alphaA-conotoxins EIVA and EIVB, block both Torpedo and mouse alpha1-containing muscle subtype nAChRs expressed in Xenopus oocytes at low nanomolar concentrations. In contrast to alpha-bungarotoxin, alphaA-EIVA is inactive at alpha7-containing nAChRs even at micromolar concentrations. In this regard, alphaA-EIVA is similar to the previously described alpha-conotoxins (e.g. alpha-MI and alpha-GI) which also selectively target alpha1- versus alpha7-containing nAChRs. However, alpha-MI and alpha-GI discriminate between the alpha/delta versus alpha/gamma subunit interfaces of the mouse muscle nAChR with 10,000-fold selectivity. In contrast, alphaA-conotoxin EIVA blocks both the alpha/gamma site and alpha/delta site with equally high affinity but with distinct kinetics. The alphaA-conotoxins thus represent novel probes for the alpha/gamma as well as the alpha/delta binding sites of the nAChR.

Amino Acid Sequence↗

PubMed1997

Cathepsin D displays in vitro beta-secretase-like specificity.

The formation of A beta and A beta-containing fragments is likely a key event in the process of neural degeneration in Alzheimer's disease. The N-terminal residue (Asp-1) of A beta and its C-terminally extended sequences is liberated from the beta-amyloid precursor protein (beta APP) by beta-secretase(s). This activity appears highly increased by the presence (N-terminally to Asp-1) of a double-mutation (KM-->NL) found in several Swedish families affected by early onset Alzheimer's disease. By means of synthetic peptides encompassing the "normal' (N peptide) and mutated (delta NL peptide) sequences targeted by beta-secretase(s), we have detected a human brain protease displaying preferred efficiency for the delta NL peptide than for the non-mutated analog. This activity is sensitive to pepstatin, maximally active at acidic pH and hydrolyses the two peptides at the expected M/D or L/D cleavage sites. Such acidic activity is also detected in rat brain, PC12 cells and primary cultured astrocytes. The pepstatin sensitivity and pH maximum of the brain activity that appeared reminiscent of those displayed by the acidic protease cathepsin D led us to examine this enzyme as a putative beta-secretase-like candidate. Purified cathepsin D displays higher catalytic parameters for the delta NL peptide than for the non-mutated peptide, cleaves these two substrates at the expected M/D or L/D sites, and is maximally active at acidic pH. However, cathepsin D does not cleave peptides bearing mutations that were previously shown to drastically lower or fully block A beta secretion by transfected cells. Furthermore, cathepsin D hydrolyses recombinant baculoviral delta NL beta APP751 at a 6-fold higher rate than beta APP751 and gives rise to a 12-kDa C-terminal product that is recognized by antibodies fully specific of the N-terminus of A beta. Altogether, our study indicates that cathepsin D displays several in vitro beta-secretase-like properties that suggests that this protease could fulfill such a role, at least in the Swedish genetic form of Alzheimer's disease.

Alzheimer Disease↗

PubMed1997

Colocalization of tissue transglutaminase and stress fibers in human vascular smooth muscle cells and human umbilical vein endothelial cells.

The subcellular distribution of tissue transglutaminase in human umbilical vein endothelial cells and human arterial and venous smooth muscle cells was examined. Double-immunofluorescence staining of smooth muscle cells and endothelial cells with anti-transglutaminase antisera and rhodamine-tagged phalloidin revealed codistribution of transglutaminase with the stress fibers, with endothelial cells also containing a cytoplasmic pool. This pattern of distribution was confirmed by confocal microscopy. Immunoprecipitation experiments demonstrated that transglutaminase co-immunoprecipitated with myosin in high-molecular-weight complexes, but not with actin, suggesting that the association of transglutaminase with the stress fibers was due to its cross-linking to myosin. About 97% of endothelial cell transglutaminase activity was present in the cytosolic fraction and 3% in the particulate fraction. The detergent-insoluble fraction was practically devoid of activity as measured by the putrescine assay, but was active as evidenced by the covalent cross-linking of 125I-fibronectin. Western blotting with a polyclonal rabbit antiserum raised against human erythrocyte transglutaminase detected high levels of enzyme in endothelial cell cytosol and both detergent-soluble and detergent-insoluble membrane fractions. In contrast, smooth muscle cells contained much less cytosolic transglutaminase, as determined either functionally or antigenically. Furthermore, within the particulate fraction of the smooth muscle cells, most of the enzyme was located in the detergent-insoluble fraction, as assessed by Western blot analysis. Retinoic acid increased the levels of enzyme in the cytosol of all cell types and the increases were correlated with increases in mRNA. Thus, tissue transglutaminase is present in various particulate fractions of vascular smooth muscle cells and endothelial cells and may be present in this cellular fraction by virtue of autocross-linking of the enzyme itself to stress fiber-associated myosin.

Blotting, Northern↗

PubMed1997

Identification of two amino acids of the human cholecystokinin-A receptor that interact with the N-terminal moiety of cholecystokinin.

A region between residues 38 and 42 of the human cholecystokinin-A (CCK-A) receptor was shown to be involved in the binding of CCK but not in that of JMV 179 and JMV 180, two peptides closely related to CCK (Kennedy, K., Escrieut, C., Dufresne, M., Clerc, P., Vaysse, N., and Fourmy, D. (1995) Biochem. Biophys. Res. Commun. 213, 845-852). In the present study, we have identified the residues of both the receptor and the ligand responsible for this differential binding. Residues Trp-39 and Gln-40 of the receptor were crucial for binding of the C-terminal nonapeptide of CCK as W39F and Q40N mutants demonstrated parallel decreases in both affinity and potency to induce accumulation of inositol phosphates (12.9- and 20.9-fold). The W39F and Q40N mutant receptors bound CCK analogues modified at their C-terminal end, including JMV 179 and JMV 180, as well as the C-terminal amidated heptapeptide of CCK, with identical affinities to the wild-type receptor. In contrast, W39F and Q40N mutants bound CCK octapeptide with the same decreased affinity as the CCK nonapeptide. The modeling of the CCK-A receptor and the docking of the peptide agonists [Thr,Nle]CCK9 and CCK-8 indicated that their N terminus was connected to the receptor through a strong bond network involving Trp-39 and Gln-40 thus confirming experimental data. These first molecular data identifying the agonist binding site of the human CCK-A receptor represent an important step toward the complete delineation of the agonist binding site and the understanding of the molecular mechanisms that govern differential activation of this receptor by CCK-related peptides.

Amino Acid Sequence↗

PubMed1997

Tissue-specific inhibition of apolipoprotein B mRNA editing in the liver by adenovirus-mediated transfer of a dominant negative mutant APOBEC-1 leads to increased low density lipoprotein in mice.

APOBEC-1 is a catalytic subunit of an apolipoprotein B (apoB) mRNA editing enzyme complex. In humans it is expressed only in the intestine, whereas in mice it is expressed in both the liver and intestine. APOBEC-1 exists as a spontaneous homodimer (Lau, P. P., Zhu, H.-J., Baldini, A., Charnsangavej, C., and Chan, L. (1994) Proc. Natl. Acad. Sci. U. S. A. 91, 8522-8526). We tested the editing activity and dimerization potential of three different mouse APOBEC-1 mutants using in vitro editing activity assay and immunoprecipitation in the presence of epitope-tagged APOBEC-1. One catalytically inactive mutant, mu1 (H61K/C93S/C96S), that retains its capacity to dimerize with wild-type APOBEC-1 was found to inhibit the editing activity of the latter and was thus a dominant negative mutant. Two other inactive mutants that dimerized poorly with APOBEC-1 failed to inhibit its activity. Intravenous injection of a mu1 adenovirus, Admu1, in C57BL/6J mice in vivo resulted in liver-specific expression of mu1 mRNA. On days 4 and 9 after virus injection, endogenous hepatic apoB mRNA editing was 23.3 +/- 5.0 and 36.8 +/- 5.7%, respectively, compared with 65.3 +/- 11 and 71.3 +/- 5.2%, respectively, for luciferase adenovirus-treated animals. Plasma apoB-100 accounted for 95 and 93% of total plasma apoB in Admu1 animals on days 4 and 9, respectively, compared with 78 and 72% in luciferase adenovirus animals. Plasma cholesterol on day 9 was 98 +/- 17 mg/dl in the mu1-treated animals, substantially higher than phosphate-buffered saline-treated (57 +/- 9 mg/dl) or luciferase-treated (71 +/- 12 mg/dl) controls. Fast protein liquid chromatography analysis of mouse plasma showed that the intermediate density/low density lipoprotein fractions in the animals treated with the dominant negative mutant adenovirus were much higher than those in controls. We conclude that active APOBEC-1 functions as a dimer and its activity is inhibited by a dominant negative mutant. Furthermore, apoB mRNA editing determines the availability of apoB-100, which in turn limits the amount of intermediate density/low density lipoprotein that can be formed in mice. Liver-specific inhibition of apoB mRNA editing is an important component of any strategy to enhance the value of mice as a model for human lipoprotein metabolism.

APOBEC-1 Deaminase↗