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Biomedical subjects

J M Iversen

Publications and source records attributed to J M Iversen.

15 recordsLinked to original sources
PubMed2001

Parotid salivary basic proline-rich proteins inhibit HIV-I infectivity.

OBJECTIVE: The objective of this study was to investigate the molecular nature, spectrum of activity and mechanism(s) of action of those human parotid basic proline-rich proteins that exhibit anti-HIV-I activity. DESIGN: Fractions containing the basic proline-rich proteins were obtained from human parotid saliva of presumed HIV-I non-infected human subjects and characterized with respect to their purity, apparent molecular size and their ability to inhibit the infectivity of T-tropic and M-tropic strains of HIV-I. SUBJECTS, MATERIALS AND METHODS: Stimulated parotid saliva samples were collected from human subjects who denied having any risk factors for HIV-I infection and whose parotid salivas inhibited HIV-I infectivity. Such samples were subjected to affinity, molecular sieve and ion exchange chromatography to isolate individual salivary components. Those fractions demonstrating anti-HIV-I activity were analyzed by SDS-PAGE in order to assess their purity and determine their apparent molecular weights. HIV-I inhibitory activity was determined using HIV-I strains LAI and BaL in a Hela cell-derived multinuclear activation of a galactosidase indicator (MAGI) assay. Amino acid analyses were performed on some fractions. RESULTS: Recombinant gp120-CH-Sepharose chromatography of one subject's parotid saliva revealed specific binding of human parotid basic proline-rich proteins, most prominently one with an apparent molecular weight of 37 kDa. Molecular sieve and cation exchange chromatography yielded a fraction greatly enriched in this protein which amino acid analysis confirmed was proline-rich. A similar fraction from two other subjects also contained basic proline-rich proteins of similar molecular size. These fractions inhibited both T-tropic and M-tropic strains of HIV-I when assayed in the MAGI system. Since SLPI activity is not observable in the MAGI assay, this inhibition was not due to SLPI. The presence of thrombospondin-I (TSP-I) in the active fractions was precluded on the basis of SDS-PAGE examination. CONCLUSIONS: Specific basic proline-rich proteins in human parotid saliva possess significant anti-HIV-I activity independent of that attributable to SLPI or TSP-I. Since the inhibition is detectable with the MAGI assay, its mechanism of action involves virus-host cell interaction prior to the introduction of the tat gene product into the host cell and may be through the binding of the basic proline-rich proteins to the HIV-I gp120 coat of the virus.

Amino Acids↗
PubMed1994

Oligonucleotide probes for mutans streptococci.

Oligodeoxyribonucleotide probes complementary to hypervariable regions of the 16S ribosomal RNA were designed for specificity toward Streptococcus mutans and Streptococcus sobrinus. The probes were tested for specificity and sensitivity by hybridization with nucleic acid from over 100 mutans and non-mutans oral streptococci and other common oropharyngeal bacteria. Probes designated SM002 and SM010 were 100% sensitive and > 99% specific for S. mutans. Probe SSP001, designed to detect both S. mutans and S. sobrinus, was 88% sensitive and > 99% specific. The probes were able to detect nucleic acid extracted from 3 x 10(4)-1 x 10(5) homologous bacteria. Sensitivity did not vary significantly with the growth state of the cells, except for diminished signals when using nucleic acid extracted from very old cultures. The probes correctly identified 72 S. mutans colonies isolated from 10 volunteer saliva samples, using sugar utilization patterns as a reference standard. Ten isolates resembling S. mutans by colony morphology but not by sugar utilization patterns were correctly distinguished from mutans streptococci by the probes. These results demonstrate that oligonucleotide probes can accurately identify S. mutans in saliva samples.

Base Sequence↗
PubMed1993

Anti-infectivity activity of human salivary secretions toward human immunodeficiency virus.

The purpose of this investigation was to adapt an MT-2 cell syncytium-forming assay for measuring anti-infectivity activity of salivary secretions toward HIV and to determine the distribution of this activity in a population of healthy adult subjects. Whole saliva samples were collected from 27 volunteers, who reported that they did not belong to any group at high risk for HIV infection, and tested for anti-infectivity activity using the syncytium-forming assay. Nine of these subjects were subsequently retested on one or more occasions to assess the variability in appearance of this activity. Parotid and extraparotid salivas of six subjects were also tested. Samples were frozen immediately after collection and submitted in blinded fashion for quantitation of their anti-HIV activity using a syncytia-forming MT-2 cell assay or the p24 antigen ELISA. Nine out of the 27 subjects showed detectable anti-HIV infectivity activity. One parotid sample and one extraparotid sample out of four from subjects with positive whole salivas were positive and none of the parotid or extraparotid samples from two subjects with negative whole salivas were positive. The inhibitory activity ranged from 0.5 to 1 log 10 TCID50/ml and could not be correlated with total protein content in saliva or any specific electrophoretic component. Filtration of the saliva through an Amicon 10 filter before incubation with the virus abolished the activity. Similar studies using two other biological fluids, urine and cerebrospinal fluid, revealed no anti-HIV infectivity activity. These findings confirm the presence in saliva of inhibitory activity directed toward HIV.

Adult↗
PubMed1993

The B1-immunoreactive proteins of the perinatal submandibular gland: similarity to the major parotid gland protein, RPSP.

The B1-immunoreactive proteins of type III cells of the perinatal rat submandibular gland are immunologically cross-reactive with proteins of both the sublingual and parotid glands; in particular, protein SMG-A appears similar to a major parotid protein. We isolated SMG-A and the parotid protein (known as M1 or leucine-rich protein), prepared polyclonal antibodies to them, and compared their biochemical properties and immunological reactivities. They were identical in their molecular weight on SDS-PAGE (23.5 kDa), tenacious binding to Affi-gel Blue, isoelectric point (pH 4.53), and proteolysis to a 14 kDa peptide: Antibodies to SMG-A showed reactivity with protein SMG-C, a product of the neonatal type I cells, as well as with proteins SMG-B1 and SMG-B2, contrasted with the absence of reactivity of anti-M1 IgG with these proteins. Anti-M1 reacted with the "parotid secretory protein" (PSP) of the mouse, and M1 appears to be the homologue, in the rat, of mouse PSP.

Animals↗
PubMed1992

Use of adaptive control with feedback to individualize suramin dosing.

Suramin is the first putative growth factor inhibitor in clinical trial that has demonstrated antitumor activity. Administration of suramin is complicated by a narrow therapeutic index and significant interpatient variability of measured pharmacokinetic parameters. Because both antitumor response and dose-limiting toxicities are related to plasma suramin concentration profiles, individualized dose schedules are required for optimal administration of the compound. In this report, the use of optimal sampling theory to derive sparse data monitoring and control strategies for use with suramin is described. A fixed rate continuous infusion schedule was used in seven patients, and the time to peak concentration (280-300 micrograms/ml) ranged from 7.7-21 days (mean, 13.2 days) with a decline to 150 micrograms/ml in 3-22 days (mean, 11 days). An initial population pharmacokinetic model was fit using a maximum likelihood algorithm. The mean volume of the central compartment was 4.5 +/- 6.7 liters/m2, volume of the peripheral compartment 10.6 +/- 1.4 liters/m2, distributional half-life 25 +/- 5.4 h, and elimination half-life 29.7 +/- 6.9 h. The terminal half-life was shorter than previously reported. These parameters were used as the initial population model for an iterative 2-stage analysis. The resulting distributional half-life of 22.3 +/- 2.7 h and elimination half-life of 28.2 +/- 5.0 h were similar, reflecting the intensive sampling. The iterative 2-stage analysis model was then used to determine the optimal sampling times and to simulate 20 data sets for a protocol designed to maintain plasma concentrations in a defined concentration range. This strategy is currently under investigation in phase I clinical trials.

Adenocarcinoma↗
PubMed1992

Inhibition of synthesis of rat parotid secretory proteoglycan in a gland slice system.

The chondroitin sulphate contained within the secretory granules of the rat parotid gland and its saliva was shown to be in the form of a proteoglycan by using inhibitors of proteoglycan synthesis in a gland slice system. Gland slices were incubated in either p-nitrophenyl-beta-D-xyloside or chlorate in the presence of both [3H]-leucine and [35S]-sulphate. The slices were next homogenized and either the 250 g supernatant fraction (for initial experiments) or secretory granule-containing fractions were isolated. Protein and proteoglycans of these fractions were precipitated in 10% trichloracetic acid (TCA), and glycosaminoglycans in cetylpyridinium chloride. [3H]-leucine and [35S]-sulphate were quantitated in each type of precipitate by scintillation counting. The results showed that 1 mM xyloside had no effect on protein or glycosaminoglycan synthesis but blocked incorporation of radiosulphate into TCA-precipitable material. Sixteen mM chlorate almost totally inhibited incorporation of radiosulphate into glycosaminoglycan and TCA-precipitable material. These findings demonstrate that the rat parotid secretory chondroitin sulphate is indeed a proteoglycan because its synthesis is blocked by the protein-core analogue acceptor, p-nitrophenyl-beta-D-xyloside. This system offers opportunities for exploring the functional role of chondroitin sulphate proteoglycan in this salivary gland.

Animals↗
PubMed1992

Therapeutic alternatives for hormone-refractory prostatic cancer.

The currently available prognostic factors allow the identification of patients who are destined to do poorly with primary hormone therapy. Standardization of these factors is required so that they become incorporated into routine practice. For patients who relapse in osseous sites radiolabeled diphosphonates are one therapeutic alternative that can provide palliation. Future use will be directed to optimize treatment schedules, and use earlier in the course of the disease so that more durable palliation of bony metastases can be achieved. Evolving data on the use of PSA show that elevations antedate clinical relapse for most patients. In these cases, sequential changes can be used to assess therapeutic effects, which in turn can allow novel therapies to be screened more rapidly in the clinic. Criteria for the degree of change that is indicative of response must be standardized. Preliminary data suggest a 50% decline be the minimum that is used. The results with agents such as suramin, which interrupts autocrine and paracrine growth factor loops, are a therapeutic strategy that need further study. Future efforts will focus on defining which patients are best suited for specific therapeutic approaches.

Antigens, Neoplasm↗
PubMed1991

Maturation-related changes in mass and elemental contents of secretory granules as measured by electron-microprobe.

The relationship between granule density, protein content, and Ca and S contents were studied in two secretory granule fractions, from parotid glands of the rat, previously shown to constitute different stages in granule maturation. The density of the lighter fraction was between 1.133 and 1.142 g/ml, while that of the heavier fraction was greater than 1.142 g/ml. The mean protein content of the denser granules was 12% greater than that of the lighter granules (P less than 0.03), while the dry-mass elemental concentrations in the two granule fractions were unchanged. These results indicate that protein is added to granules during the maturation process (presumably by vesicular traffic), and that the resulting increase in granule density is not driven simply by decrease in water content and/or increased concentrations of inorganic Ca or S in the granules. The elemental concentration values also indicate that the diffusible elements permeate the granule membrane during the fractionation procedures.

Animals↗
PubMed1991

Microprobe analysis of maturation-related elemental changes in rat parotid secretory granules.

Electron probe X-ray microanalysis was use to quantitate the elemental and mass changes that take place during the secretory granule maturation process. A single injection of isoproterenol stimulated the depletion of secretory granules from rat parotid acinar cells. Granules at different stages of maturation were analyzed as they reaccumulated within the cells over time. Dry mass measurements revealed that secretory material becomes concentrated about twofold within maturing granules. Nearly all of the increase in mass concentration could be attributed to a reduction in water space. Data are presented that indicate that Na, K, Cl, and water all efflux from secretory granules during maturation. In contrast, granule S content is positively correlated with maturation. Hence, significant changes in granule elemental and water contents occur during the maturation process.

Animals↗
PubMed1987

The presence of chondroitin sulfate in parotid secretory granules and saliva of the rat.

The presence of chondroitin sulfate in secretory granules of the rat parotid gland and its saliva was revealed by radioactive sulfate incorporation, followed by isolation and partial characterization of the sulfated species contained within the granules and in the parotid saliva. 35SO4 was incorporated into chromatographically identical macromolecular material both in vitro, in a gland-slice system followed by isolation of granule contents, and in vivo as measured in the pure parotid secretion following intravenous administration of 35SO4=. The majority of the 35SO4= label appeared in a peak in the region where the family of acidic proline-rich proteins elute from a DEAE-Sephadex A-50 column. Papain digestion freed the sulfated material from the bulk of protein present in this peak, leaving sulfate-labelled material that chromatographed on Sepharose CL6B as a single peak corresponding to a molecular weight of 13,000 daltons. The ratio of uronic acid to amino sugar in this sulfated peak was 0.56. The sulfated material was susceptible to degradation by chondroitinase AC. The presence of this chondroitin sulfate in secretory granules and saliva is consistent with previous suggestions that sulfated polyanions may play a role in formation and maturation of secretory granules.

Animals↗
PubMed1985

Isolation and partial characterization of two populations of secretory granules from rat parotid glands.

A method is described for the isolation of two populations of secretory granules from rat parotid glands utilizing differences in their sedimentation characteristics. The granule preparations were analyzed for homogeneity by electron microscopy and chemical analyses. The soluble contents of both types of granules were obtained by hypotonic lysis, and the proteins compared by SDS-PAGE and ion exchange-gel filtration chromatography. Both populations of secretory granules appear to have the same protein composition as that of the parotid saliva. The secretory granules with the smaller apparent buoyant density became labelled with radioactive leucine earlier than the heavier granules when a pulse of this amino acid was supplied to a gland slice system. The lighter granules appear to represent an earlier stage in maturation.

Animals↗
PubMed1982

Isolation and characterization of the basic proline-rich proteins from rat parotid saliva.

Five fractions of basic proline-rich proteins were isolated from rat parotid saliva, obtained by surgical cannulation of the ducts. The purification procedures employed DEAE-Sephadex to isolate a heterogeneous break-through fraction containing the basic proline-rich proteins, followed by gel filtration on Sephadex G-200 to separate the high molecular weight glycoprotein, fraction A, from the other basic proline-rich proteins which were resolved into four additional fractions, SP-1 to SP-4, by ion exchange chromatography on SP-Sephadex. The proteins differed in their amino acid composition and content of neutral and amino sugars. All the proteins were characterized by a high proportion of proline (approx. 40 mol per cent) and glycine (11-23 mol per cent). Four of the fractions were also enriched in glutamic acid/glutamine (19-26 mol per cent). The exception was fraction SP-4, which contained lower levels of glutamic acid/glutamine and has no counterpart in human basic proline-rich proteins. Fraction A, the basic glycoprotein, was heavily glycosylated (59 mol per cent), whereas SP-2 and SP-4 were less glycosylated. Fractions SP-1 and SP-3 contained low levels of neutral and amino sugars. Basic proline-rich proteins constitute a smaller percentage of the total protein in rat parotid saliva than they do in human parotid saliva (10.5 versus 40 per cent). Rat basic glycoprotein fraction constitutes less than 1 per cent whereas the human glycoprotein fraction constitutes 17 per cent. Rat basic proline-rich proteins appear to be larger and less basic than most of the human basic proteins, and they resolve into fewer protein fractions (4 versus 9) with SP-Sephadex chromatography.

Amino Acids↗
PubMed1980

The presence of mast cell granules in rat parotid secretory granule preparations.

Rat parotid secretory granule preparations contain, in addition to the acinar secretory granules, a second type of granule. Whereas the acinar granules lyse under hypotonic conditions, this second type of granule does not, thus providing a means for obtaining a fraction sufficiently enriched in these granules to allow for their characterization. In the present study, these granules are shown to possess demonstrable chymotrypsin-like enzyme activity. In the intact rat parotid, such activity is shown by histochemical methods to be present in the numerous mast cells residing in the connective tissue stroma, but no such activity exists in any of the parenchymal cells. On the basis of their electron microscopic appearance, enzyme activity, and physical characteristics it is concluded that the second type of granule present in rat parotid secretory granule preparations originates from stromal mast cells rather than from parenchymal cells.

Animals↗
PubMed1977

Changes in rat parotid salivary proteins induced by chronic isoproterenol administration.

Changes in the rat parotid gland and its secretion, brought about by chronic isoproterenol administration, were studied. In addition to the expected enlargement, morphological and biochemical analyses of the glands showed evidence of changes in the secretory components. Chromatographic and electrophoretic experiments revealed both qualitative and quantitative changes in the secretory proteins.

Amino Acids↗