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J P Henry

Publications and source records attributed to J P Henry.

At least 19 recordsLinked to original sources
PubMed1992

Characterization and purification of the monoamine transporter of bovine chromaffin granules.

The monoamine transporter of the chromaffin granule membranes can be specifically labeled by the photoaffinity reagent 7-azido-8-[125I]iodoketanserin. The characteristics of the labeled protein have been investigated. Two-dimensional gel electrophoresis of the labeled membranes indicated a MW of about 70,000 and an isoelectric point ranging from 3.8 to 4.6. No clear protein spot was associated with the radioactive material, which migrated between glycoproteins GPII and GPIV. The diffuse aspect of the radioactive material indicated a heterogeneity, which was not modified after a second electrophoresis. This heterogeneity was, at least partially, due to glycosylation of the transporter; neuraminidase treatment increased the protein pI up to 6.3, whereas digestion with N-glycopeptidase markedly decreased the apparent MW, from 70,000 to 50,000. SDS-polyacrylamide gel electrophoresis showed that, at low acrylamide concentrations, the labeled material migrated more rapidly than predicted from the mobility of the markers of molecular weight, a behavior which indicated a marked hydrophobicity of the transporter. The labeled protein was purified to homogeneity by a combination of chromatography on DEAE-cellulose at pH 4.5, on immobilized wheat germ agglutinin, and on hydroxylapatite in the presence of SDS. During this purification, the specific radioactivity was increased by a factor of 300-500, with a yield of 10-20%.

Affinity Labels↗
PubMed1992

Properties of the mitochondrial peptide-sensitive cationic channel studied in planar bilayers and patches of giant liposomes.

A voltage-dependent cationic channel of large conductance is observed in phospholipid bilayers formed by the tip-dip method from proteoliposomes derived from mitochondrial membranes. It is blocked by peptide M, a 13 residue peptide having the properties of a mitochondrial signal sequence. To verify the reliability of the experimental approach, mitochondrial membranes from bovine adrenal cortex or porin-deficient mutant yeast were either fused to planar bilayers or incorporated in giant liposomes which were studied by patch clamp. Cationic channels were found with both techniques. They had the same conductance levels and voltage-dependence as those which have been described using the tip-dip method. Moreover, they were similarly blocked by peptide M. The voltage-dependence of block duration was analyzed in planar bilayer and tip-dip records. Results strengthen the idea that peptide M might cross the channel. Other mitochondrial channels were observed in planar bilayers and patch clamp of giant liposomes. Because they were never detected in tip-dip records, they are likely to be inactivated at the surface monolayer used to form the bilayer in this type of experiment.

Adrenal Cortex↗
PubMed1991

Use of spin-labeled and fluorescent lipids to study the activity of the phospholipid transfer protein from maize seedlings.

The transfer of spin-labeled and fluorescent lipids between sonicated vesicles and different host membranes has been measured in the presence or absence of a phospholipid transfer protein purified from maize seedlings. It was found that the protein has little specificity towards the phospholipid head group and allows the transfer of hydrophobic long chain phospholipids. By contrast, no transfer of a cholesterol analogue could be detected. By EPR spectroscopy, evidence is presented that shows that the protein catalyzes the incorporation of labeled phospholipids in the outer monolayer of the acceptor membranes. The efficiency of the transfer depends largely on the nature of the acceptor: erythrocytes are more difficult to label than chromaffin granules or liposomes made with unsaturated lipids. Thus, consistent with the high activation energy observed, the transfer is facilitated when it involves fluid membranes. These results are in favor of a process involving the exchange of phospholipids, facilitated by a shuttle protein rather than a fusion mechanism.

Ascorbic Acid↗
PubMed1991

A peptide-sensitive channel of large conductance is localized on mitochondrial outer membrane.

Applying the technique of 'tip-dip' to mitochondria, we have shown the existence in this organelle of a cationic channel of large conductance, which is blocked by a 13-residue peptide possessing the sequence of the N-terminal extremity of the cytochrome c oxidase subunit IV precursor. To study the submitochondrial localization of the channel, the effect of trypsin on isolated channels and on entire mitochondria were compared. One side of isolated channels is sensitive to trypsin, which eliminates the voltage dependence. Channels isolated from trypsinized mitochondria were devoid of voltage dependence and were blocked by the peptide. This suggests a localization of the channel on the outer membrane. Consistent with this hypothesis, the channel was observed with the highest frequency in outer membrane fractions purified by different procedures, either from bovine adrenal cortex or from rat liver mitochondria. Such a localization is also consistent with digitonin solubilization experiments. The channel was solubilized before the inner membrane marker, cytochrome c oxidase. The orientation of the channel was inferred from its trypsin sensitivity and its potential dependence: a transmembrane potential (inside negative) will close the channel.

Animals↗
PubMed1991

A 28 kDa mitochondrial protein is radiolabelled by crosslinking with a 125I-labelled presequence.

A 13-residue peptide containing the first 12 amino acids of the N-terminal part of the signal sequence of yeast cytochrome c oxidase subunit IV is shown by chemical crosslinking to interact with a mitochondrial protein. This result is obtained with mitochondria from four different origins. Submitochondrial localization experiments suggest that the 28 kDa labelled component is present on the outer face of the inner membrane. Since such addressing peptides are imported into mitochondria through the same machinery as protein precursors, the 28 kDa protein might be a component of the translocation apparatus.

Amino Acid Sequence↗
PubMed1991

The participation of annexin II (calpactin I) in calcium-evoked exocytosis requires protein kinase C.

Permeabilized adrenal chromaffin cells secrete catecholamines by exocytosis in response to micromolar calcium concentrations. Recently, we have demonstrated that chromaffin cells permeabilized with digitonin progressively lose their capacity to secrete due to the release of certain cytosolic proteins essential for exocytosis (Sarafian T., D. Aunis, and M. F. Bader. 1987. J. Biol. Chem. 34:16671-16676). Here we show that one of the released proteins is calpactin I, a calcium-dependent phospholipid-binding protein known to promote in vitro aggregation of chromaffin granules at physiological micromolar calcium levels. The addition of calpactin I into digitonin- or streptolysin-O-permeabilized chromaffin cells with reduced secretory capacity as a result of the leakage of cytosolic proteins partially restores the calcium-dependent secretory activity. This effect is specific of calpactin I since other annexins (p32, p37, p67) do not stimulate secretion at similar or higher concentrations. Calpactin I requires the presence of Mg-ATP, suggesting that a phosphorylating step may regulate the activity of calpactin. Calpactin is unable to restore the secretory activity in cells which have completely lost their cytosolic protein kinase C or in cells having their protein kinase C inhibited by sphingosine or downregulated by long-term incubation with TPA. In contrast, calpactin I prephosphorylated in vitro by purified protein kinase C is able to reconstitute secretion in cells depleted of their protein kinase C activity. This stimulatory effect is also observed with thiophosphorylated calpactin I which is resistant to cellular phosphatases or with phosphorylated calpactin I introduced into cells in the presence of microcystin, a phosphatase inhibitor. These results suggest that calpactin I is involved in the exocytotic machinery by a mechanism which requires phosphorylation by protein kinase C.

Adenosine Triphosphate↗
PubMed1991

Amphiphilic and nonamphiphilic forms of bovine and human dopamine beta-hydroxylase.

We show that human and bovine dopamine beta-hydroxylases (DBH) exist under three main molecular forms: a soluble nonamphiphilic form and two amphiphilic forms. Sedimentation in sucrose gradients and electrophoresis under nondenaturing conditions, by comparison with acetylcholinesterase (AChE), suggest that the three forms are tetramers of the DBH catalytic subunit and bind either no detergent, one detergent micelle, or two detergent micelles. By analogy with the Gna4 and Ga4 AChE forms, we propose to call the nonamphiphilic tetramer Dna4 and the amphiphilic tetramers Da4I and Da4II. In addition to the major tetrameric forms, DBH dimers occur as very minor species, both amphiphilic and nonamphiphilic. Reduction under nondenaturing conditions leads to a partial dissociation of tetramers into dimers, retaining their amphiphilic character. This suggests that the hydrophobic domain is not linked to the subunits through disulfide bonds. The two amphiphilic tetramers are insensitive to phosphatidylinositol phospholipase C, but may be converted into soluble DBH by proteolysis in a stepwise manner; Da4II----Da4I----Dna4. Incubation of soluble DBH with various phospholipids did not produce any amphiphilic form. Several bands corresponding to the catalytic subunits of bovine DBH were observed in sodium dodecyl sulfate-polyacrylamide gel electrophoresis, but this multiplicity was not simply correlated with the amphiphilic character of the enzyme. In the case of human DBH, we observed two bands of 78 and 84 kDa. As previously reported by others, the presence of the heavy subunit characterizes the amphiphilic forms of the enzyme. We discuss the nature of the hydrophobic domain, which could be an uncleaved signal peptide, and the organization of the different amphiphilic and nonamphiphilic DBH forms. We present two models in which dimers may possess either one hydrophobic domain or two domains belonging to each subunit; in both cases, a single detergent micelle would be bound per dimer.

Acetylcholinesterase↗
PubMed1991

Single-parameter DNA flow cytometric analysis of normal-appearing colonic mucosa does not predict the presence of colonic neoplasia.

We wished to determine whether DNA flow cytometric analysis could detect DNA abnormalities in normal-appearing mucosa of patients with colonic neoplasia. Eighty-five patients were studied either at colonoscopy or at surgical resection. Forty-five had macroscopically normal colonoscopy; 13 had adenomatous polyps, and 27 had colorectal carcinoma. Biopsies were obtained from the cancer and from normal-appearing mucosa 5 cm from the lesion. The patients who had normal colonoscopy had rectal biopsies. The samples were prepared for analysis on a Coulter EPICS C flow cytometer. Cells were analyzed for presence of aneuploidy (%AN), percent in DNA synthetic phase (%S), and percent growth fraction (%GF = %S + %G2M). Aneuploidy was present in 12 of 27 carcinomas (44%), but in none of the samples from polyps or normal-appearing colorectal mucosa adjacent to cancers. The %S from cancers was greater than those from polyps (9.6 +/- 6.3 vs. 5.1 +/- 1.8, p less than 0.005). However, %S from specimens arising from normal-appearing mucosa 5 cm distant from cancer could not be differentiated from the rectal mucosa of macroscopically normal colons (5.9 +/- 2.5 vs. 5.2 +/- 2.7). The %GF of cancer specimens was greater than those from adenomas (26.0 +/- 11.0 vs. 10.8 +/- 3.7, p less than 0.005). However, the %GF of normal-appearing mucosa 5 cm distant from the cancer was similar to the findings from mucosa arising from macroscopically normal colons (10.5 +/- 3.3 vs. 11.0 +/- 3.4). In conclusion, DNA flow cytometric analysis of normal-appearing colonic mucosa from patients with colonic carcinoma does not demonstrate abnormalities of DNA content or cell cycle kinetics, and therefore, cannot predict the presence of colonic neoplasia.

Adult↗
PubMed1990

The rise of body temperature induced by the stimulation of dopamine D1 receptors is increased in acutely reserpinized mice.

In naive mice, the selective D1 agonist, SK&F 38393 (7.5-30 mg/kg s.c.), induced a significant rise of body temperature (0.5-1 degree C) which was antagonized by SCH 23390 (100 micrograms/kg s.c.) and by flupenthixol (0.4 mg/kg i.p.). In mice treated with reserpine (5 mg/kg s.c.) 18 h before testing, which on its own caused intense hypothermia (10-12 degrees C), SK&F 38393 (1.87-30 mg/kg s.c.) induced a dose-dependent and more marked rise of body temperature (5-7 degrees C). Similarly, SK&F 38393 (30 mg/kg s.c.) partially prevented reserpine-induced hypothermia. The central origin of the SK&F 38393 effects in reserpine-treated mice is indicated by the rise of body temperature induced by the i.c.v. administration of the drug (12.5-50 micrograms per mice). The SK&F 38393-induced rise of body temperature in acutely reserpinized mice was antagonized by SCH 23390 (50-200 micrograms/kg s.c.), clozapine (1.87-30 mg/kg i.p.) or chlorpromazine (2-32 mg/kg i.p.) but not by metoclopramide (25 or 100 mg/kg i.p.) or amisulpride (12.5 or 50 mg/kg). In naive mice, apomorphine (1 mg/kg s.c.) or LY 171555 (0.4 mg/kg s.c.) induced hypothermia which was antagonized by amisulpride (12.5 mg/kg i.p.); a transiently increased body temperature was even measured 30 min after apomorphine injection in amisulpride-treated mice. Apomorphine (1 mg/kg s.c.) induced a rise of body temperature in acutely reserpinized mice which was significantly reduced by SCH 23390 (50 and 200 micrograms/kg s.c.) and significantly increased by amisulpride (12.5 and 50 mg/kg i.p.). These data suggest that pharmacologically different dopamine receptor subtypes mediate different effects on body temperature in mice: D1 dopamine receptors mediate a rise of body temperature which is increased in hypothermic reserpinized animals and dopamine receptors of the D4 subtype mediate the decrease of body temperature in naive mice.

2,3,4,5-Tetrahydro-7,8-dihydroxy-1-phenyl-1H-3-ben↗
PubMed1990

Subcellular distribution of acetylcholinesterase forms in chromaffin cells. Do chromaffin granules contain a specific secretory acetylcholinesterase?

The presence of acetylcholinesterase (AChE) in chromaffin granules has been controversial for a long time. We therefore undertook a study of AChE molecular forms in chromaffin cells and of their distribution during subcellular fractionation. We characterized four main AChE forms, three amphiphilic forms (Ga1, Ga2 and Ga4), and one non-amphiphilic form (Gna4). Each form shows the same molecular characteristics (sedimentation, electrophoretic migration, lectin interactions) in the different subcellular fractions. All forms are glycosylated and seem to possess both N-linked and O-linked carbohydrate chains. There are differences in the structure of the glycans carried by the different forms, as indicated by their interaction with some lectins. Glycophosphatidylinositol-specific phospholipases C converted the Ga2 form, but not the other amphiphilic forms, into non-amphiphilic derivatives. The distinct patterns of AChE molecular forms observed in various subcellular compartments indicate the existence of an active sorting process. Gna4 was concentrated in fractions of high density, containing chromaffin granules. We obtained evidence for the existence of a lighter fraction also containing chromogranin A, tetrabenazine-binding sites and Gna4 AChE, which may correspond to immature, incompletely loaded granules or to partially emptied granules. The distribution of Gna4 during subcellular fractionation suggested that this form is largely, but not exclusively, contained in chromaffin granules, the membranes of which may contain low levels of the three amphiphilic forms.

Acetylcholinesterase↗
PubMed1990

Comparison of mitochondrial cationic channels in wild-type and porin-deficient mutant yeast.

Bilayers were formed at the tip of microelectrodes from a suspension of proteoliposomes derived from wild-type and porin-deficient mutant yeast mitochondria. In both preparations, identical cationic channels of large conductance were recorded. This result rules out any relationship between this channel and the outer membrane voltage-dependent anion channel, the activity of which is carried by porin. The ionic selectivity and the voltage-dependence of the yeast cationic channel suggest that it is related to that recently described in mammalian mitochondria. This hypothesis is further supported by the fact that both channels are blocked by a mitochondrial addressing peptide.

Amino Acid Sequence↗
PubMed1990

Comparative evaluation of the diagnosis of acute pancreatitis based on serum and urine enzyme assays.

We compared the diagnostic sensitivities of serum amylase, lipase (assayed enzymatically and immunologically), trypsinogen and elastase-1, the 2-h-timed urine amylase excretion and the ratio of amylase and creatinine clearances in the recognition of acute pancreatitis. Serial serum and urine findings from 39 patients with acute pancreatitis, and from 42 patients with non-pancreatic causes of abdominal pain (controls), as well as findings from 24 healthy subjects (normals) were studied. Decision thresholds were established for each parameter using either the control or the normal population, and the resulting diagnostic sensitivities determined. On hospital admission, all serum assays were equally sensitive, but on subsequent days lipase, trypsinogen and elastase-1 assays all significantly surpassed the sensitivity of the serum amylase assay. On the second and subsequent hospitalization days, determination of timed urine amylase excretion offered no advantage over the serum amylase, and the ratio of amylase and creatinine clearances lacked discrimination altogether.

Acute Disease↗
PubMed1990

Atypical eating disorder masquerading as recurrent acute pancreatitis: the value of multiple pancreatic serological markers.

A 28-year-old woman with nausea, vomiting, and abdominal pain had been hospitalized elsewhere on 13 separate occasions over the year before this admission for similar episodes thought to be secondary to acute pancreatitis. She had undergone repeated work-ups including endoscopic retrograde cholangiopancreatography, computed tomographic scan, and exploratory laparotomy. There was a discrepancy between her unremarkable physical examination and extremely elevated amylase (3,210 U/L) which suggested nonpancreatic hyperamylasemia; normal serum pancreatic isoamylase, trypsinogen, and lipase confirmed this suspicion. The patient was noted to have self-induced vomiting in the hospital which she admitted was frequent behavior. her psychiatric disturbance was characterized as an atypical eating disorder. This case illustrates that hyperamylasemia in association with abdominal pain, nausea, and vomiting may not be secondary to pancreatitis and that use of a second serum marker (such as trypsinogen, lipase, or isoamylase) helps to establish a definitive diagnosis.

Acute Disease↗
PubMed1990

[Comparison of 24-hour gastric pH in normal controls and patients with duodenal ulcer or atrophic gastritis].

Twenty-four intragastric acidity was measured by continuous recording using intragastric glass electrodes in 16 normal controls, 18 inactive duodenal ulcer patients and 7 patients with atrophic gastritis. Median pH for the 24 h period, for the 8 AM-8 PM period, and for the 8 PM-8 AM period were significantly lower in duodenal ulcer patients than in controls (1.19 vs 1.78; 1.17 vs 2.05; 1.27 vs 1.64). Median pH for these 3 periods were significantly higher in patients with atrophic gastritis than in controls (3.90; 3.72; 3.81). The median pH for the 24 h period was higher than the lower quartile value of the control group in 33 per cent of duodenal ulcer patients. During the night (24 h-3 h), duodenal ulcer patients had significantly lower median pH (1.03) than controls (1.51). Antisecretory treatment should be directed to decrease this period of unbuffered acidity.

Adult↗