Biomedical subjects

P Chen

Publications and source records attributed to P Chen.

At least 235 records · Page 13Linked to original sources

PubMed1999

Atrial natriuretic-like peptide and its prohormone within metasequoia.

Metasequoia glyptostroboides was one of the dominant conifers in North America, Asia, and Europe for more than 100 million years since the late Cretaceous Albian Age, but Quaternary glaciations drove the Metasequoia population to apparent extinction. A small pocket of Metasequoia, however, was found in central China in the 1940s representing the only surviving population of this "living fossil" species. Atrial natriuretic peptide, a 28-amino-acid peptide hormone that causes sodium and water excretion in animals, has been found to be part of the first peptide hormonal system in lower plants. The existence of this hormonal system has never been examined within trees of any genus. High-performance gel permeation chromatography of the leaves and stems (i.e., branches) of Metasequoia followed by atrial natriuretic peptide radioimmunoassay revealed an ANP-like peptide and its prohormone (i.e., approximately 13,000 mol wt) were present in both leaves and stems of this conifer. The elution profile of ANP-like peptide in stems of Metasequoia had a shoulder to the left of where pure synthetic ANP elutes suggesting the possibility of a slightly larger peptide eluting within this shoulder secondary to alternate processing of the ANP-like prohormone and similar to what occurs with the kidney of animals. The elution profile of ANP-like peptide in the leaves of Metasequoia revealed two peaks; one where ANP elutes and a second peak suggesting a smaller peptide that has been metabolically processed. The presence of the ANP-like prohormone strongly suggests that ANP-like gene expression is occurring in both leaves and stems of Metasequoia since this prohormone is the gene product of this hormonal system. The presence of the ANP-like hormonal system in trees implies that this hormonal system may have been present early in land plant evolution to allow trees to reach heights of greater than 30 feet where a water flow-enhancing substance is absolutely necessary for water flow to occur to these heights.

Atrial Natriuretic Factor↗

PubMed1999

Structural characterization of the N-linked oligosaccharides in bile salt-stimulated lipase originated from human breast milk.

The detailed structures of N- glycans derived from bile salt-stimulated lipase (BSSL) found in human milk were determined by combining exoglycosidase digestion with matrix-assisted laser desorption/ionization time-of-flight mass spectrometry. The N- glycan structures were conclusively determined in terms of complexity and degree of fucosylation. Ion-exchange chromatography with pulsed amperometric detection, together with mass-spectral analysis of the esterified N- glycans, indicated the presence of monosialylated structures. The molecular mass profile of esterified N- glycans present in BSSL further permitted the more detailed studies through collision-induced dissociation (CID) and sequential exoglycosidase cleavages. The N- glycan structures were elucidated to be complex/dibranched, fucosylated/complex/dibranched, monosialylated/complex/dibranched, and monosialylated/fucosylated/dibranched entities.

Carbohydrate Sequence↗

PubMed1999

Stable elliptical vortices in a circular disk.

We show that large elliptical vortices in a finite disk are stable in a two-dimensional (2D) ideal fluid (this also applies to a column of quasi-2D non-neutral plasma in an axial magnetic field). The stability is established by comparison between the energy of elliptical and symmetrical states to satisfy a sufficient condition, without dynamical eigenanalysis. Analytical small ellipticity expansion of system energy and exact numerical values for arbitrary ellipticity are both obtained for uniform vortices. An approximating calculation is presented for general smooth vortices. Numerical simulations of the 2D Euler equation are also performed. The simulations not only confirm the sufficient condition, but also show that the stability persists to smaller vortex sizes. The reason why decaying l=2 modes were obtained by Briggs, Daugherty, and Levy [Phys. Fluids 13, 421 (1970)] using eigenanalysis is also discussed.

Journal Article↗

PubMed1999

Amplitude equation and pattern selection in Faraday waves.

A nonlinear theory of pattern selection in parametric surface waves (Faraday waves) is presented that is not restricted to small viscous dissipation. By using a multiple scale asymptotic expansion near threshold, a standing wave amplitude equation is derived from the governing equations. The amplitude equation is of gradient form, and the coefficients of the associated Lyapunov function are computed for regular patterns of various symmetries as a function of a viscous damping parameter gamma. For gamma approximately 1, the selected wave pattern comprises a single standing wave (stripe pattern). For gamma<<1, patterns of square symmetry are obtained in the capillary regime (large frequencies). At lower frequencies (the mixed gravity-capillary regime), a sequence of sixfold (hexagonal), eightfold, ...patterns are predicted. For even lower frequencies (gravity waves) a stripe pattern is again selected. Our predictions of the stability regions of the various patterns are in quantitative agreement with recent experiments conducted in large aspect ratio systems.

Journal Article↗

PubMed1999

Functional analyses of the promoters in the lantibiotic mutacin II biosynthetic locus in Streptococcus mutans.

The lantibiotic bacteriocin mutacin II is produced by the group II Streptococcus mutans. The mutacin II biosynthetic locus consists of seven genes, mutR, -A, -M, -T, -F, -E, and -G, organized as two operons. The mutAMTFEG operon is transcribed from the mutA promoter 55 bp upstream of the translation start codon for MutA, while the mutR promoter is 76 bp upstream of the mutR structural gene. Expression of the mutA promoter is regulated by the components of the growth medium, while the mutR promoter activity does not seem to be affected by these conditions. Inactivation of mutR abolishes transcription of the mutA operon but does not affect its own promoter activity. The expressions of both mutA and mutR promoters are independent of the growth stage, while the production of mutacin II is only elevated at the early stationary phase. Taken together, these results suggest that expression of the mutacin operon is regulated by a complex system involving transcriptional and posttranscriptional or posttranslational controls.

Bacteriocins↗

PubMed1999

The specific genes for lantibiotic mutacin II biosynthesis in Streptococcus mutans T8 are clustered and can be transferred en bloc.

Mutacin II is a ribosomally synthesized peptide lantibiotic produced by group II Streptococcus mutans. DNA sequencing has revealed that the mutacin II biosynthetic gene cluster consists of seven specific open reading frames: a regulator (mutR), the prepromutacin structural gene (mutA), a modifying protein (mutM), an ABC transporter (mutT), and an immunity cluster (mutFEG). Transformations of a non-mutacin-producing strain, S. mutans UA159, and a mutacin I-producing strain, S. mutans UA140, with chromosomal DNA from S. mutans T8 with an aphIII marker inserted upstream of the mutacin II structural gene yielded transformants producing mutacin II and mutacins I and II, respectively.

Bacteriocins↗

PubMed1999

Purification of mutacin III from group III Streptococcus mutans UA787 and genetic analyses of mutacin III biosynthesis genes.

Previously, members of our group reported the isolation and characterization of mutacin II from Streptococcus mutans T8 and the genetic analyses of the mutacin II biosynthesis genes (J. Novak, P. W. Caufield, and E. J. Miller, J. Bacteriol. 176:4316-4320, 1994; F. Qi, P. Chen, and P. W. Caufield, Appl. Environ. Microbiol. 65:652-658, 1999; P. Chen, F. Qi, J. Novak, and P. W. Caufield, Appl. Environ. Microbiol. 65:1356-1360, 1999). In this study, we cloned and sequenced the mutacin III biosynthesis gene locus from a group III strain of S. mutans, UA787. DNA sequence analysis revealed eight open reading frames, which we designated mutR, -A, -A', -B, -C, -D, -P, and -T. MutR bears strong homology with MutR of mutacin II, while MutA, -B, -C, -D, -P, and -T are counterparts of proteins in the lantibiotic epidermin group. MutA' has 60% amino acid identity with MutA and therefore appears to be a duplicate of MutA. Insertional inactivation demonstrated that mutA is an essential gene for mutacin III production, while mutA' is not required. Mutacin III was purified to homogeneity by using reverse-phase high-pressure liquid chromatography. N-terminal peptide sequencing of the purified mutacin III determined mutA to be the structural gene for prepromutacin III. The molecular mass of the purified peptide was measured by laser disorption mass spectrophotometry and found to be 2,266.43 Da, consistent with our supposition that mutacin III has posttranslational modifications similar to those of the lantibiotic epidermin.

Amino Acid Sequence↗

PubMed1999

Virulence of Mycobacterium tuberculosis CDC1551 and H37Rv in rabbits evaluated by Lurie's pulmonary tubercle count method.

The virulence of the CDC1551 strain of Mycobacterium tuberculosis was compared to that of H37Rv in a rabbit inhalation model. While rabbits that inhaled the two strains produced equal numbers of grossly visible primary tubercles, CDC1551 tubercles were smaller and contained fewer bacilli than H37Rv tubercles. These findings suggest that a miniepidemic near the Kentucky-Tennessee border caused by CDC1551 was due not to increased virulence but to increased transmissibility.

Animals↗

PubMed1999

Conditional sigma factor expression, using the inducible acetamidase promoter, reveals that the Mycobacterium tuberculosis sigF gene modulates expression of the 16-kilodalton alpha-crystallin homologue.

A chemically inducible acetamidase promoter-sigF fusion gene was integrated into the chromosome of Mycobacterium bovis BCG. Two-dimensional protein gel analysis permitted the identification of a number of protein spots whose expression was SigF related. One spot upregulated by inappropriate induction of sigF expression corresponded to the 16-kDa antigen alpha-crystallin.

Amidohydrolases↗

PubMed1999

Endothelin-1 enhances plasminogen activator inhibitor-1 production by human brain endothelial cells via protein kinase C-dependent pathway.

The effects of endothelin-1 (ET-1) on the production of plasminogen activator inhibitor 1 (PAI-1) and tissue plasminogen activator (t-PA) by human brain-derived endothelial cells in culture were studied. At 100 nmol/L, ET-1 increased PAI-1 production by 88+/-6% within 72 hours, and increased PAI-1 mRNA expression within 1 hour of stimulation; there was no significant effect on t-PA production. PAI-1 activity was also examined and found to increase with ET-1 treatment. Suboptimal concentrations of ET-1 and tumor necrosis factor-alpha (TNF-alpha) acted synergistically to increase PAI-1 production. ET-1 activated protein kinase C and cAMP-dependent protein kinase pathways within 3 to 5 minutes of treatment, with the peak at 10 minutes. Activation of protein kinase C by phorbol-12-myristate-13-acetate (PMA) resulted in increased PAI-1 production, whereas activation of the cAMP-dependent protein kinase by forskolin or dibutyryl cAMP (dBu-cAMP) significantly decreased PAI-1 production. However, simultaneous activation of protein kinase C by PMA and cAMP-dependent protein kinase by dBu-cAMP only slightly attenuated PMA-induced PAI-1 increase. Inhibition of protein kinase C by GF-109213X abolished the effects of ET-1. These results demonstrate that ET-1 and TNF-alpha function synergistically to induce procoagulant activity of brain endothelial cells in a process that involves a protein kinase C-dependent pathway.

Brain↗

PubMed1999

Nicotine increases plasminogen activator inhibitor-1 production by human brain endothelial cells via protein kinase C-associated pathway.

BACKGROUND AND PURPOSE: Smoking both increases stroke risk and reduces the risk of thrombolysis-associated intracerebral hemorrhage. Plasminogen activator inhibitor-1 (PAI-1) is a major regulator of fibrinolysis; elevation of PAI-1 is associated with an increased risk of thrombotic disorders. We studied the effect of nicotine, an important constituent of cigarette smoke, on PAI-1 production by human brain endothelial cells. METHODS: Adult human central nervous system endothelial cells (CNS-EC) were used for tissue culture experiments. We analyzed culture supernatant for PAI-1 protein and measured PAI-1 mRNA (by Northern blot analysis) and protein kinase C (PK-C) activity. RESULTS: Nicotine at 100 nmol/L increased PAI-1 protein production and mRNA expression by CNS-EC. After 72 hours of exposure to nicotine, the concentration of secreted PAI-1 in the cell supernatant was increased 1.90+/-0.2 fold compared with untreated cells. PAI-1 mRNA also increased approximately twofold. Inhibition of PK-C completely abolished this effect. Nicotine had no effect on the concentration of tissue plasminogen activator. CONCLUSIONS: Nicotine increases brain endothelial cell PAI-1 mRNA expression and protein production via PK-C-dependent pathway. These findings provide new insights into why smoking may be associated with predisposition to thrombosis and inversely associated with intracerebral hemorrhage after therapeutic tissue plasminogen activator therapy.

Adult↗

PubMed1999

Neuropeptide Y and tuberoinfundibular dopamine activities are altered during lactation: role of prolactin.

During lactation the suckling stimulus increases the activity of two populations of neuropeptide Y (NPY) neurons in the hypothalamus, the caudal portion of the arcuate nucleus (ARH) and the dorsomedial hypothalamus (DMH), and suppresses the activity of TIDA neurons in the ARH. In the present study, an acute resuckling model was used to examine the role of suckling-induced hyperprolactinemia in modulating the activity of these systems. Lactating rats were deprived of their eight-pup litters on day 9 postpartum, and 48 h later, the animals served either as nonsuckled controls (0 pups) or were suckled for 24 h. In addition, some of the resuckled animals received two s.c. injections of bromocriptine (0.5 mg/rat x injection), a dopamine D2 agonist, to inhibit suckling-induced PRL secretion. In situ hybridization was performed for rat NPY messenger RNA (mRNA) and tyrosine hydroxylase (TH) mRNA to provide an index for NPY and TIDA neuronal activities, respectively. Resuckling for 24 h induced a significant increase in NPY mRNA levels in the caudal portion of the ARH and in the DMH. Bromocriptine treatment did not alter the increase in NPY mRNA levels in the ARH, whereas the treatment greatly attenuated the increase in NPY mRNA in the DMH. TH mRNA levels in the rostral ARH area returned to basal levels in the nonsuckled control animals, and 24 h of resuckling significantly suppressed TH mRNA expression in this area. Bromocriptine treatment caused a significant increase in TH mRNA levels compared with those in the eight-pup suckled group. Thus, the results from the present study demonstrate that the suckling stimulus activated the two populations of NPY neurons and suppressed TIDA activity. Suckling-induced hyperprolactinemia did not participate in the increase in ARH NPY activity, whereas it played a major stimulatory role in suckling-induced activation of NPY neurons in the DMH and an inhibitory role in suckling-induced suppression of TIDA activity. The increase in TIDA activity after bromocriptine treatment was unexpected and suggests that the role of PRL in the regulation of TIDA activity is significantly altered during lactation.

Animals↗

PubMed1999

Morphological evidence for direct interaction between arcuate nucleus neuropeptide Y (NPY) neurons and gonadotropin-releasing hormone neurons and the possible involvement of NPY Y1 receptors.

Neuropeptide Y (NPY) neurons in the arcuate nucleus of the hypothalamus (ARH) have been shown to play an important role in modulating LH secretion. One mechanism by which the ARH NPY system may regulate LH secretion is by modulating GnRH neuronal function. Thus, the present study examined whether the ARH NPY system provided direct input to GnRH cell bodies in the preoptic area (POA), as well as to their nerve terminals in the median eminence (ME). The possible involvement of the NPY Y1 receptor subtype in mediating the effects of NPY was also investigated. Lactating rats were used in these studies because they have increased hypothalamic NPY content, especially in the ARH/ME areas, making it easier to detect NPY fibers and terminals. The anterograde tracer, Phaseolus vulgaris leucoagglutinin (PHA-L), was iontophoresed into the ARH of lactating rats; and triple-label immunofluorescence was performed, with the aid of confocal microscopy, to visualize NPY, PHA-L, and GnRH. GnRH cell bodies were found scattered throughout the organum vasculosum laminae terminalis (OVLT)/POA region, and NPY/ PHA-L double-labeled fibers were found in very close proximity to numerous GnRH perikarya. In the ME, double-labeled NPY/PHA-L fibers were found in the inner and external zones, and they were found in close proximity to GnRH neuronal fibers. Using a NPY Y1 specific antibody, double-label immunofluorescence was performed to examine whether the Y1 receptor subtype was expressed in GnRH neurons. No convincing Y1-positive staining was found in GnRH cell bodies in the OVLT/POA region. However, abundant Y1-positive fiber and cell staining were observed throughout the region, and Y1-positive fibers were found in close apposition to GnRH cell bodies. In contrast, numerous GnRH nerve fibers and terminals in both the OVLT and ME were colocalized with Y1-positive staining. The results of this study suggest that ARH NPY neurons come in close contact with GnRH neurons and may provide direct input to both GnRH cell bodies in the POA region and to their nerve terminals in the ME. The Y1 receptor subtype may be directly involved in NPY modulation of GnRH secretion from its nerve terminals.

Animals↗

PubMed1999

Characterization of the neuroanatomical distribution of agouti-related protein immunoreactivity in the rhesus monkey and the rat.

Agouti-related protein (AGRP) is a recently described homolog of the skin agouti protein. AGRP is transcribed primarily in the adrenal and hypothalamus and is a high affinity antagonist of the neural melanocortin-3 and melanocortin-4 receptors. The perikarya expressing AGRP messenger RNA are found in the arcuate nucleus of the rat and rhesus monkey. Using a polyclonal antibody against the pharmacologically active domain of AGRP (amino acids 83-132), we have also characterized the distribution of AGRP-immunoreactive neurons in both species. The major fiber tracts are conserved in both species, with dense projections originating in the arcuate nucleus and proceeding along the third ventricle. Dense fiber bundles are also visible in the paraventricular, dorsomedial, and posterior nuclei in the hypothalamus, in the bed nucleus of the stria terminalis, and in the lateral septal nucleus of the septal region. AGRP-containing neurons are not visualized in a number of areas, including portions of the amygdala, thalamus, and brain stem, that express MC3-R and MC4-R messenger RNA and receive innervation from POMC neurons that serve as the source of melanocortin agonists. Thus, AGRP is most likely to be involved in modulating a conserved subset of the physiological functions of central melanocortin peptides. Based on the particular distribution of AGRP neurons, those functions are likely to include the central control of energy homeostasis.

Animals↗

PubMed1999

Altered expression of agouti-related protein and its colocalization with neuropeptide Y in the arcuate nucleus of the hypothalamus during lactation.

During lactation, the levels of neuropeptide Y (NPY), which plays an important role in mediating food intake, are significantly elevated in a number of hypothalamic areas, including the arcuate nucleus (ARH). To identify additional hypothalamic systems that might be important in mediating the increase in food intake and alterations in energy homeostasis during lactation, the present studies examined the expression of agouti-related protein (AGRP), a recently described homologue of the skin agouti protein. AGRP is found in the hypothalamus and has been suggested to play an important role in the regulation of food intake. In the first experiment, animals were studied during diestrus of the estrous cycle, a stage of the cycle when estrogen levels are basal and similar to lactation, or during days 12-13 postpartum. Lactating animals had their litters adjusted to eight pups on day 2 postpartum. Brain tissue sections were used to measure AGRP messenger RNA (mRNA) levels by in situ hybridization. AGRP mRNA signal was found mostly in the ventromedial portion of the ARH, which has been shown to contain a high density of NPY neurons. A significant increase in AGRP mRNA content was observed in the mid- to caudal portion of the ARH of lactating animals compared with diestrous females. No difference was found in the rostral portion of the ARH. In the second experiment, double-label in situ hybridization for AGRP and NPY was performed in lactating animals to determine the extent of colocalization of the two peptides in the ARH, using 35S-labeled and digoxigenin-labeled antisense complementary RNA probes. It was found that almost all of the NPY-positive neurons throughout the ARH also expressed AGRP mRNA signal. Furthermore, AGRP expression was confined almost exclusively to NPY-positive neurons. Thus, the present study showed that during lactation, AGRP gene expression was significantly elevated in a subset of the AGRP neurons in the ARH. The high degree of colocalization of AGRP and NPY, coupled with previous reports from our laboratory demonstrating increased NPY expression in the ARH in response to suckling, suggests that AGRP and NPY are coordinately regulated and may be involved in the increase in food intake during lactation.

Agouti Signaling Protein↗

PubMed1999

Mechanisms of GDF-5 action during skeletal development.

Mutations in GDF-5, a member of the TGF-beta superfamily, result in the autosomal recessive syndromes brachypod (bp) in mice and Hunter-Thompson and Grebe-type chondrodysplasias in humans. These syndromes are all characterised by the shortening of the appendicular skeleton and loss or abnormal development of some joints. To investigate how GDF-5 controls skeletogenesis, we overexpressed GDF-5 during chick limb development using the retrovirus, RCASBP. This resulted in up to a 37.5% increase in length of the skeletal elements, which was predominantly due to an increase in the number of chondrocytes. By injecting virus at different stages of development, we show that GDF-5 can increase both the size of the early cartilage condensation and the later developing skeletal element. Using in vitro micromass cultures as a model system to study the early steps of chondrogenesis, we show that GDF-5 increases chondrogenesis in a dose-dependent manner. We did not detect changes in proliferation. However, cell suspension cultures showed that GDF-5 might act at these stages by increasing cell adhesion, a critical determinant of early chondrogenesis. In contrast, pulse labelling experiments of GDF-5-infected limbs showed that at later stages of skeletal development GDF-5 can increase proliferation of chondrocytes. Thus, here we show two mechanisms of how GDF-5 may control different stages of skeletogenesis. Finally, our data show that levels of GDF-5 expression/activity are important in controlling the size of skeletal elements and provides a possible explanation for the variation in the severity of skeletal defects resulting from mutations in GDF-5.

Amino Acid Sequence↗

PubMed1999

p27(Kip1) links cell proliferation to morphogenesis in the developing organ of Corti.

Strict control of cellular proliferation is required to shape the complex structures of the developing embryo. The organ of Corti, the auditory neuroepithelium of the inner ear in mammals, consists of two types of terminally differentiated mechanosensory hair cells and at least four types of supporting cells arrayed precisely along the length of the spiral cochlea. In mice, the progenitors of greater than 80% of both hair cells and supporting cells undergo their terminal division between embryonic day 13 (E13) and E14. As in humans, these cells persist in a non-proliferative state throughout the adult life of the animal. Here we report that the correct timing of cell cycle withdrawal in the developing organ of Corti requires p27(Kip1), a cyclin-dependent kinase inhibitor that functions as an inhibitor of cell cycle progression. p27(Kip1) expression is induced in the primordial organ of Corti between E12 and E14, correlating with the cessation of cell division of the progenitors of the hair cells and supporting cells. In wild-type animals, p27(Kip1) expression is downregulated during subsequent hair cell differentiation, but it persists at high levels in differentiated supporting cells of the mature organ of Corti. In mice with a targeted deletion of the p27(Kip1) gene, proliferation of the sensory cell progenitors continues after E14, leading to the appearance of supernumerary hair cells and supporting cells. In the absence of p27(Kip1), mitotically active cells are still observed in the organ of Corti of postnatal day 6 animals, suggesting that the persistence of p27(Kip1) expression in mature supporting cells may contribute to the maintenance of quiescence in this tissue and, possibly, to its inability to regenerate. Homozygous mutant mice are severely hearing impaired. Thus, p27(Kip1) provides a link between developmental control of cell proliferation and the morphological development of the inner ear.

Animals↗

PubMed1999

Rapid Screening of Olefin Polymerization Catalyst Libraries by Electrospray Ionization Tandem Mass Spectrometry.

The simultaneous screening of catalysts according to their propensity for catalyzing the polymerization of ethylene (see scheme) can be achieved with the help of electrospray ionization tandem mass spectrometry. A small (eight catalysts) library of Pd(II) complexes synthesized simultaneously in a one-pot reaction has demonstrated the efficiency of this new screening technique. This method could be widened to libraries of other olefin polymerization catalysts.

Journal Article↗