Mechanisms of transcriptional activation by retinoic acid receptors.
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Animals↗
Biomedical subjects
L Xu
Publications and source records attributed to L Xu.
Adenosine triphosphate activates a noninactivating K+ current in adrenal cortical cells through nonhydrolytic binding.
Bovine adrenal zona fasciculata (AZF) cells express a noninactivating K+ current (IAC) that is inhibited by adrenocorticotropic hormone and angiotensin II at subnanomolar concentrations. Since IAC appears to set the membrane potential of AZF cells, these channels may function critically in coupling peptide receptors to membrane depolarization, Ca2+ entry, and cortisol secretion. IAC channel activity may be tightly linked to the metabolic state of the cell. In whole cell patch clamp recordings, MgATP applied intracellularly through the patch electrode at concentrations above 1 mM dramatically enhanced the expression of IAC K+ current. The maximum IAC current density varied from a low of 8.45 +/- 2.74 pA/pF (n = 17) to a high of 109.2 +/- 26. 3 pA/pF (n = 6) at pipette MgATP concentrations of 0.1 and 10 mM, respectively. In the presence of 5 mM MgATP, IAC K+ channels were tonically active over a wide range of membrane potentials, and voltage-dependent open probability increased by only approximately 30% between -40 and +40 mV. ATP (5 mM) in the absence of Mg2+ and the nonhydrolyzable ATP analog AMP-PNP (5 mM) were also effective at enhancing the expression of IAC, from a control value of 3.7 +/- 0.1 pA/pF (n = 3) to maximum values of 48.5 +/- 9.8 pA/pF (n = 11) and 67.3 +/- 23.2 pA/pF (n = 6), respectively. At the single channel level, the unitary IAC current amplitude did not vary with the ATP concentration or substitution with AMP-PNP. In addition to ATP and AMP-PNP, a number of other nucleotides including GTP, UTP, GDP, and UDP all increased the outwardly rectifying IAC current with an apparent order of effectiveness: MgATP > ATP = AMP-PNP > GTP = UTP > ADP >> GDP > AMP and ATP-gamma-S. Although ATP, GTP, and UTP all enhanced IAC amplitude with similar effectiveness, inhibition of IAC by ACTH (200 pM) occurred only in the presence of ATP. As little as 50 microM MgATP restored complete inhibition of IAC, which had been activated by 5 mM UTP. Although the opening of IAC channels may require only ATP binding, its inhibition by ACTH appears to involve a mechanism other than hydrolysis of this nucleotide. These findings describe a novel form of K+ channel modulation by which IAC channels are activated through the nonhydrolytic binding of ATP. Because they are activated rather than inhibited by ATP binding, IAC K+ channels may represent a distinctive new variety of K+ channel. The combined features of IAC channels that allow it to sense and respond to changing ATP levels and to set the resting potential of AZF cells, suggest a mechanism where membrane potential, Ca2+ entry, and cortisol secretion could be tightly coupled to the metabolic state of the cell through the activity of IAC K+ channels.
Adenosine Diphosphate↗
Use of retroviral vectors encoding murine inducible nitric oxide synthase gene to suppress tumorigenicity and cancer metastasis of murine melanoma.
The purpose of this study was to determine whether retrovirus-mediated transfer of murine macrophage inducible nitric oxide synthase (iNOS) can produce inhibition of tumorigenicity and metastasis. Retroviral vectors encoding macrophage iNOS constructed in pLXSN, a retroviral vector with the iNOS gene under the control of a long terminal repeat promoter, were stably transfected into PA317 cells. Medium harvested from confluent monolayers of the virus-producing cell lines was used for infection of the murine K-1735 melanoma cells. Expression of iNOS was confirmed by northern and Western blot analyses. Functional iNOS protein expression was confirmed by bioassay of nitrite accumulation in the culture supernatant. Cells infected by a control iNOS-negative retrovirus produced fast-growing subcutaneous tumors and many lung metastases in nude mice, whereas iNOS-transduced cells produced slow-growing tumors and few lung metastases, showing that the infection of murine tumor cells by retroviruses harboring the iNOS gene can suppress tumorigenicity and metastasis.
Animals↗
Comparison of quantitative HCV RNA assays in chronic hepatitis C.
We compared the relative sensitivities of first-and-second generation branched nucleotide assays (Quantiplex HCV RNA 1.0 and 2.0, respectively, Chiron, Emeryville, Calif) for the detection of hepatitis C virus (HCV) RNA to that of a commercially available quantitative reverse transcriptase polymerase chain reaction (RT-PCR) method (Monitor, Roche Molecular Systems, Nutley, NJ) in 53 patients with chronic hepatitis C. The sensitivities of the second-generation branched DNA (bDNA) and RT-PCR assays were similar (91% and 92%, respectively), and both were significantly more sensitive (P < .001) than the first-generation method. Moreover, both assays detected HCV RNA in all 11 patients with type 2a, 2b, or 3a genotypes vs 45% with the HCV RNA 1.0 bDNA assay. We examined 174 serum samples by the bDNA 2.0 and RT-PCR assays. Major quantification differences were noted on a given specimen with the RT-PCR method reporting values an average 41-fold lower (range, 0-703-fold) than those obtained with the bDNA assay. We conclude that both methods can be used to detect HCV RNA in patients who are infected with the genotypes that are most commonly encountered in the United States. The HCV RNA 2.0 bDNA assay may offer advantages when attempting to quantify high-level viremia.
Chronic Disease↗
Cellular responses to surgical trauma, hemorrhage, and resuscitation with diaspirin cross-linked hemoglobin in rats.
BACKGROUND: Resuscitation with acellular oxygen carrier solutions offers the potential advantage of improved oxygen delivery compared with crystalloid solutions, but the detailed consequences of improved resuscitation have not been fully evaluated. This study evaluated local and systemic cellular effects of trauma, hemorrhage, and resuscitation in a model of hemorrhage and surgical trauma. METHODS: Rats with a 10 cm full-thickness incisional wound and a 15 mL/kg hemorrhage were either not resuscitated or resuscitated with blood or diaspirin cross-linked hemoglobin (DCLHb). Cellular proliferative responses were evaluated at 1.5, 6, 24, and 48 hours after wounding by labeling in vivo with 5-bromo-2'-deoxyuridine. Plasma levels of interleukin-6, tumor necrosis factor-alpha, and interferon-gamma were measured by bioassay or enzyme-linked immunosorbent assay (ELISA). Bacterial translocation was measured by culturing liver homogenates. RESULTS: Trauma inhibited keratinocyte and hepatocyte proliferation at 1.5 and 6 hours, and stimulated subsequent proliferation of keratinocytes and liver nonparenchymal cells. DCLHb stimulated wound keratinocyte proliferation, attenuated the inhibition of hepatocyte proliferation, eliminated bacterial translocation to the liver, protected the intestine from ischemic damage, and induced a rapid increase of interleukin-6 during the early phase of injury. CONCLUSIONS: Surgical trauma alone, or in combination with hemorrhage, modulated cell proliferation both in the wound and in the remote organs of intestine and liver. DCLHb enhanced wound healing and cell proliferation as well as, or better than, freshly drawn blood, which may be beneficial for trauma care.
Animals↗
Promoter analysis and expression of a phospholipase D gene from castor bean.
The expression of a castor bean (Ricinus communis L.) phospholipase D (PLD; EC 3.1.4.4) gene has been studied by examining its promoter activity in transgenic tobacco (Nicotiana tabacum) carrying a PLD promoter-glucuronidase transgene and by monitoring the levels of PLD mRNA in castor bean. Sequence and the 5' truncation analyses revealed that the 5' flanking region from nucleotide -1200 to -730 is required for the regulation and basal function of the PLD promoter. The PLD promoter in vegetative tissues is highly active in the rapidly growing regions such as the shoot apex and the secondary meristem producing axillary buds and vascular tissues of young leaves and stems. The PLD promoter activity in floral tissues was high in stigma, ovary, and pollen grains, but low in petals, sepals, the epidermis of anthers, styles, and filaments. The PLD promoter activity was enhanced by abscisic acid. Northern-blot analysis of PLD in castor bean showed that the PLD mRNA levels were high in young and metabolically more active tissues such as expanding leaves, hypocotyl hooks, developing seeds, and young seedlings, and they decreased in mature tissues such as fully expanded leaves and developed seeds. These patterns of expression suggest a role of PLD in rapid cell growth, proliferation, and reproduction.
Base Sequence↗
Transfection with aFGF cDNA improves wound healing.
Somatic gene therapy is a potentially useful strategy for the delivery of growth factors or cytokines to enhance wound healing. Experimental excisional and incisional wounds in impaired-healing diabetic mice (db/db) were treated with aFGF and with a plasmid coding for aFGF. A eukaryotic expression plasmid composed of the Hst signal peptide sequence in-frame with the human aFGF sequence was used. Transfection of tissues was accomplished either by direct plasmid uptake or by uptake facilitated with cationic liposomes. The results show that the closure of excisional wounds was significantly accelerated (p < 0.05) by topical application of human recombinant aFGF or by transfection with the aFGF plasmid but not by vehicle or control plasmid not containing the aFGF sequence. In incisional wounds, aFGF or transfection with the plasmid significantly increased the wound-breaking strength compared to their corresponding controls (p < 0.05). Quantitative histology of the plasmid-treated incisional wound sections revealed improved wound quality. The transcription of mRNA from human aFGF cDNA in the incisional wound tissue extracts was confirmed by RT-PCR, and the expressed aFGF was detected by immune dot blot and immunohistochemistry assays. The transfection was a transient process with a peak at 9 d in db/+ (littermates of the diabetic mice) incisional wounds, at 36 d in db/db incisional wounds, and at 27 d in db/db excisional wounds. Cells transfected with human aFGF occupied up to 6.4% of the transectional area in the wound sites. Thus, aFGF gene delivery resulted in both gene expression and a functional improvement in healing.
Animals↗
Immunohistological study of lesions induced by Porphyromonas gingivalis in a murine model.
A previous study used a mouse model to demonstrate protection after challenge with Porphyromonas gingivalis ATCC 33277. In the present study, this same model was used to determine the phenotype of cells recruited into the lesions during the course of the protective immune response after immunization with this periodontal pathogen. BALB/c mice were immunized with 100 micrograms of P. gingivalis outer membrane antigens per mouse weekly for 3 weeks followed by challenge with live organisms 3 weeks after the final immunization. Hematoxylin and eosin-stained sections showed inflammatory infiltrates in all lesions from control (immunized with adjuvant only) and immunized mice. The lesions developed central necrotic cores surrounded by neutrophils, phagocytic macrophages and lymphocytes. Neutrophils were the predominant cells in the lesions 1 day after challenge with significantly more in immunized than control mice. Acid phosphatase and nonspecific esterase-positive macrophages were detected at day 4 and became the predominant cells in the healing lesions. CD4- and CD8-positive T-cells were present from day 1, and while numbers increased over time, there were no significant differences in control or immunized mice. When mice were depleted of CD4 or CD8 cells prior to immunization with P. gingivalis, fewer neutrophils were found in the lesions 1 day after challenge compared with undepleted immunized mice. Acid phosphatase and nonspecific esterase-positive macrophages were not affected by T-cell depletion. The results suggest that the P. gingivalis-induced lesion in immunized BALB/c mice is consistent with a strong innate immune response involving the recruitment of neutrophils in the first instance which may be under the control of T cells. This is followed by the infiltration of phagocytic macrophages which are involved in the healing process and do not appear to be regulated by T cells.
Analysis of Variance↗
Cell and viral regulatory elements enhance the expression and function of a human immunodeficiency virus inhibitory gene.
Regulated expression of recombinant genes in CD4+ cells is an important objective for gene therapy of AIDS, as these cells represent the principal target for viral replication of human immunodeficiency virus (HIV). We report here that specific combinations of CD4 cell-specific and viral regulatory elements can enhance expression of an antiviral gene product. Different viral regulatory elements were incorporated into a previously reported CD4 locus control region to increase the expression of reporter genes in T and monocytic cell lines. The CD4-specific regulatory elements were included to enhance expression in CD4 cells, and viral regulatory regions, including the cytomegalovirus immediate-early (CMV IE) upstream enhancer, which contains the kappa B and Ap1 regulatory elements and a Tat-responsive element of the HIV type 1 long terminal repeat, were used to increase gene expression and modulate its activity in response to viral infection. In transient transfection assays, this vector was 100- to 1,000-fold more active than the original CD4 regulatory elements alone. Expression of an inhibitory form of the Rev protein, Rev M10, was more effective than previously described vectors and protected against productive viral replication in CD4+ peripheral blood mononuclear cells. The combination of CD4 lineage-specific and viral regulatory elements will facilitate the development of more effective antiviral genetic strategies for AIDS.
CD4 Antigens↗
Assignment of SATB1 to human chromosome band 3p23 by in situ hybridization.
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Chromosome Mapping↗
Assignment of mitotic arrest deficient protein 2 (MAD2L1) to human chromosome band 5q23.3 by in situ hybridization.
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Calcium-Binding Proteins↗
Sodium intake, angiotensin II receptor blockade, and baroreflex function in conscious rats.
The hypothesis that endogenous angiotensin II (Ang II) chronically supports baroreflex control of lumbar sympathetic nerve activity (LSNA) and heart rate (HR) via AT1 but not AT2 receptors was tested in conscious, normotensive rats. Rats were fed either a sodium deficient diet (LS) to increase circulating Ang II or a high-sodium diet (HS) for 2 to 3 weeks. One to two days after surgery to implant catheters and nerve electrodes, baroreflex curves were produced before and 40 minutes after intravenous administration of the AT1 antagonist losartan (10 mg/kg) or the AT2 antagonist PD123319 (500 micrograms/kg + 50 micrograms.kg-1.min-1). Mean arterial pressure (MAP) after losartan was maintained at basal levels with methoxamine. Forty minutes after losartan in LS rats, LSNA (46 +/- 5 to 22 +/- 1% max) and HR (414 +/- 7 to 387 +/- 8 bpm) were decreased (P < .05). Losartan decreased reflex control of LSNA more in LS than in HS rats (P < .05), as indicated by reductions in maximum LSNA (98 +/- 2 to 78 +/- 3% max) and minimum LSNA (42 +/- 5 to 21 +/- 5% max). Losartan also shifted reflex control of LSNA to a lower pressure in both groups, but the effect was larger in LS rats (-21 +/- 3 [LS] versus -9 +/- 2 [HS] mm Hg at basal LSNA; P < .05). Maximum gain was unaltered in either group. Similarly, losartan reduced maximum HR (534 +/- 6 to 495 +/- 9 bpm) and shifted the HR curve leftward (114 +/- 5 to 105 +/- 4 mm Hg) in LS but not in HS rats. In general, no changes were observed in MAP or baroreflex control of LSNA and HR after PD123319 in LS rats. These results suggest that in conscious, normotensive LS rats, endogenous Ang II supports LSNA and HR over a wide MAP range via AT1 but not AT2 receptors.
Angiotensin Receptor Antagonists↗
Topography of ribosomes and initiation complexes from rat liver as revealed by atomic force microscopy.
Atomic force microscopy (AFM) was used to image ribosomes and ribosomal subunits (60S, 40S and native 40S ribosomal subunits) isolated from rat liver. A variety of topographic images were obtained directly and found to be consistent with models established by other biophysical methods. In addition, the ternary complex of eIF-2 x GTP x Met-tRNA(i) and the 43S preinitiation complex have been discerned by AFM directly. Detailed information about the binding sites for eIF-1A, eIF-2, eIF-3, and Met-tRNA(i) on the 40S ribosomal subunit was derived from the AFM images. Finally, factors which may give rise to artifactual images, namely, convolution of the AFM tip on ribosomes, surface tension collapse effect and dehydration, are discussed. This work demonstrates that AFM is useful for imaging ribosomes and translational complexes and provides valuable information that can be used to complement other well-established techniques.
Animals↗
Prolactin-like protein-B: heterologous expression and characterization of placental and decidual species.
Prolactin-like protein-B (PLP-B) is a member of a family of proteins expressed by the rat placenta and/or decidua with characteristics similar to prolactin (PRL). In this report, we present the heterologous expression and characterization of PLP-B. Recombinant PLP-B heterologously expressed in Chinese hamster ovary cells exhibited similar immunoreactive and electrophoretic characteristics with PLP-B produced by rat placental and decidual tissues. N-terminal sequencing verified the identity and purity of the recombinant PLP-B species and the site of cleavage of the signal peptide from the mature secreted PLP-B species. Polyclonal antibodies were generated to the recombinant PLP-B and used for Western blot and immunocytochemical analyses. Recombinant and native PLP-B migrated as a doublet at 30-31 kDa in SDS-PAGE under reducing conditions. Treatment of recombinant and native PLP-B with N-glycanase accelerated their electrophoretic mobility, indicative of their glycoprotein nature. PLP-B was localized exclusively to decidual cells in the developing deciduum and spongiotrophoblast cells in the placental junctional zone. The level of PLP-B protein expression dramatically declined prior to parturition. Potential PRL-like biological actions of PLP-B were also investigated. PLP-B bound weakly to ovarian and liver PRL receptors and did not stimulate the proliferation of lactogen-dependent Nb2 lymphoma cells. In conclusion, recombinant PLP-B possesses characteristics similar to native decidual and placental PLP-B and may represent a hormone/cytokine that has important modulatory actions during the establishment of pregnancy and the initiation of parturition.
Animals↗
[Mechanical ventilation therapy with permissive hypercapnia on ARDS].
OBJECTIVE: Study of mechanical ventilation (MV) therapy of ARDS (acute respiratory distress syndrome). METHODS: Ten cases of ARDS were observed. Depending on patients' kidney compensation for respiratory acidosis, low tidal volume (VT, mean = 6.5 ml/kg) was used and certain respiratory acidosis was permitted in order to decrease airway plateau pressure and lung barotrauma. On the premise of keeping patients' PaO2 about 7.3 kPa (1 kPa = 7.5 mmHg), lowest possible FiO2 (mean = 0.51) and PEEP (mean = 0.92 kPa, 1 kPa = 10.2 cmH2O) were used. RESULTS: Seven of the ten survived. Three of them had respiratory acidosis and two had lung barotrauma during MV. CONCLUSION: Using lower VT and permitting certain respiratory acidosis are worthy to be considered in MV therapy of ARDS.
Adult↗
Isolation and characterization of a new chromosome 8q24.1 band-specific microsatellite polymorphism.
A clone, containing a 24-dinucleotide (CA)-repeat sequence, was isolated from chromosome 8q24.1 band-specific pUC19 library. It was localized to target chromosome using human-rodent hybrid cell panel. Eleven alleles were estimated from 56 unrelated individuals and the heterozygosity reaches 0.835. Linkage analysis of 11 Chinese pedigrees showed significant evidence for linkage of this new marker to three dinucleotide repeat markers at 8q23-24.1 region, with the most likely position of the marker between D8S85 and D8S199.
Alleles↗
[Changes of the contents of insulin and vasoactive intestinal peptides in the patients with primary hypertension].
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Adult↗
[Effect of yifei kangliu yin on lung cancer metastasis and immune function].
OBJECTIVE: To explore the effect of Yifei Kangliu Yin (YFKLY) on metastasis of lung cancer and immune function. METHODS: (1) Observing remote metastasis of three groups (n = 94) lung cancer patients, measuring the activity of NK cell, and the levels of OKT3, OKT4 of the patients by radioimmunoassay, and doing compared analysis. (2) Observing the effect of YFKLY on C57 mice B16 melanoma, and examining T4, T8 around the foci of cancer metastasis by immunohistochemical method. RESULTS: YFKLY significantly reduced the incidence of remote metastasis in patients and reduced the metastasis foci numbers on C57 mice of B16 melanoma. It was found that NK cell activities, OKT3 and OKT4 in peripheral blood of patients significantly increased, the infiltration of T4 lymphocytes around the foci of cancer metastasis also increased. CONCLUSION: YFKLY had inhibitory effect on the metastasis of cancer cells. It had strengthened the immune functions, aroused the anti-tumor capability.
Adenocarcinoma↗