Biomedical subjects

X J Chen

Publications and source records attributed to X J Chen.

At least 19 recordsLinked to original sources

PubMed2001

Analgesic activity and opioid receptor selectivity of stereoisomers of ohmefentanyl isothiocyanate.

Ohmefentanyl is a very potent and highly selective agonist for mu-opioid receptors. We now study analgesia, in vitro activity and opioid receptor affinity of the stereoisomers of ohmefentanyl isothiocyanate. We found that some isomers of ohmefentanyl isothiocyanate had a potent analgesic effect and that all isomers except (3R,4S,2'S)-ohmefentanyl isothiocyanate had a more potent inhibitory action on the electrically evoked contractions of mouse vas deferens than of guinea pig ileum. The inhibitory actions could be antagonized by naloxone. However, compared with the activity of the corresponding stereoisomers of ohmefentanyl, these ohmefentanyl isothiocyanates had significantly reduced analgesia and in vitro activity. They also inhibited the binding of [3H]DPDPE ([D-Pen(2),D-Pen(5)]enkephalin) and [3H]DAGO ([D-Ala(2),Mephe(4),Gly-ol(5)]enkephalin) to opioid receptors in mouse brain membranes. The inhibitory effect of stereoisomers of ohmefentanyl isothiocyanate at mu-opioid receptors was markedly lower than that of their parent compounds. The affinity of stereoisomers of ohmefentanyl isothiocyanate for delta-opioid receptors was, however, greater than or equal to that of their corresponding stereoisomers of ohmefentanyl. The results showed that the introduction of an isothiocyanato group into the phenyl ring in position-1 of ohmefentanyl reduced bioactivity and affinity to mu-opioid receptors but that the selectivity of these compounds for delta-opioid receptors was enhanced. Isomer (3R,4S,2'R)-ohmefentanyl isothiocyanate showed highest selectivity for delta-opioid receptors (K(i)(mu)/K(i)(delta)=13.6) and potent analgesic activity (ED(50)=0.25 mg/kg).

Analgesics, Opioid↗

PubMed2001

Quantitative comparison of ohmefentanyl isomers induced conditioning place preference in mice.

Differences of analgesia and withdrawal response among ohmefentanyl stereoisomers have been studied. In the present study, Quantitative comparison of reinforcing effects of ohmefentanyl stereoisomers and morphine was performed by using a conditioned place preference design in mice. Results showed that morphine and ohmefentanyl stereoisomers were able to increase significantly the time spent in the drug-paired side with respect to vehicle treated animals. A good linear correlation between doses of drugs and number of mice with place preference was found within a given dose range. On the basis of the dose-response curve analysis, ohmefentanyl stereoisomers displayed a significant difference in place preference ED50. The addictive index (analgesic ED50/place preference ED50) was used to assess the addictive potential of drugs. It was demonstrated that the addictive potential of ohmefentanyl stereoisomers did not exhibit a large difference as addictive index. Among these stereoisomers, the addictive potential of compound F9208 was markedly lower than that of morphine.

Analgesics, Opioid↗

PubMed2001

Measurements of hyperpolarized gas properties in the lung. Part III: (3)He T(1).

Hyperpolarized (3)He spin-lattice relaxation was investigated in the guinea pig lung using spectroscopy and imaging techniques with a repetitive RF pulse series. T(1) was dominated by interactions with oxygen and was used to measure the alveolar O(2) partial pressure. In animals ventilated with a mixture of 79% (3)He and 21% O(2), T(1) dropped from 19.6 sec in vivo to 14.6 sec after cardiac arrest, reflecting the termination of the intrapulmonary gas exchange. The initial difference in oxygen concentration between inspired and alveolar air, and the temporal decay during apnea were related to functional parameters. Estimates of oxygen uptake were 29 +/- 11 mL min(-1) kg(-1) under normoxic conditions, and 9.0 +/- 2.0 mL min(-1) kg(-1) under hypoxic conditions. Cardiac output was estimated to be 400 +/- 160 mL min(-1) kg(-1). The functional residual capacity derived from spirometric magnetic resonance experiments varied with body mass between 5.4 +/- 0.3 mL and 10.7 +/- 1.1 mL. Magn Reson Med 45:421-430, 2001.

Animals↗

PubMed2001

Impaired CD40-signalling in CD19-deficient mice selectively affects Th2-dependent isotype switching.

Activation of B lymphocytes involves binding of antigen to the specific receptor and signalling through several membrane coreceptors, of which CD19 has been found to play a pivotal role as a response regulator. Although previous studies in CD19 gene knockout mice have demonstrated that antibody responses to T-cell-dependent antigens are strongly impaired in the absence of this coreceptor, little is known about the consequences of CD19 deficiency for the interaction between T and B cells. Here we report that Th2 co-ordinated B-cell differentiation is selectively impaired in CD19-deficient mice in response to mucosal or systemic immunizations or following an intestinal infection with Nippostrongylus brasiliensis. Whereas immunoglobulin (Ig)G1 or IgE antibody responses were low or absent, IgG2a responses were normal. The selective defect was not caused by a poor Th2-development or interleukin (IL)-4 responsiveness in CD19-deficient mice. Rather, it was the result of an impaired Th2-B cell interaction, owing to a substantially reduced ability to signal via CD40 in CD19-deficient B cells. Thus, our study in CD19-deficient mice suggests that CD40L-CD40-interactions are more important for Th2 than for Th1 co-ordinated B-cell differentiation.

Adjuvants, Immunologic↗

PubMed2001

Dual mutations reveal interactions between components of oxidative phosphorylation in Kluyveromyces lactis.

Loss of mtDNA or mitochondrial protein synthesis cannot be tolerated by wild-type Kluyveromyces lactis. The mitochondrial function responsible for rho(0)-lethality has been identified by disruption of nuclear genes encoding electron transport and F(0)-ATP synthase components of oxidative phosphorylation. Sporulation of diploid strains heterozygous for disruptions in genes for the two components of oxidative phosphorylation results in the formation of nonviable spores inferred to contain both disruptions. Lethality of spores is thought to result from absence of a transmembrane potential, Delta Psi, across the mitochondrial inner membrane due to lack of proton pumping by the electron transport chain or reversal of F(1)F(0)-ATP synthase. Synergistic lethality, caused by disruption of nuclear genes, or rho(0)-lethality can be suppressed by the atp2.1 mutation in the beta-subunit of F(1)-ATPase. Suppression is viewed as occurring by an increased hydrolysis of ATP by mutant F(1), allowing sufficient electrogenic exchange by the translocase of ADP in the matrix for ATP in the cytosol to maintain Delta Psi. In addition, lethality of haploid strains with a disruption of AAC encoding the ADP/ATP translocase can be suppressed by atp2.1. In this case suppression is considered to occur by mutant F(1) acting in the forward direction to partially uncouple ATP production, thereby stimulating respiration and relieving detrimental hyperpolarization of the inner membrane. Participation of the ADP/ATP translocase in suppression of rho(0)-lethality is supported by the observation that disruption of AAC abolishes suppressor activity of atp2.1.

Adenosine Diphosphate↗

PubMed2001

Activity of the Kluyveromyces lactis Pdr5 multidrug transporter is modulated by the Sit4 protein phosphatase.

A possible role for posttranslational modifications in regulating the activity of ATP-binding cassette (ABC) transporters has not been well established. In this study, the drug efflux ABC transporter gene KlPDR5 was isolated from the budding yeast Kluyveromyces lactis, and it was found that the encoded KlPdr5 drug pump is posttranslationally regulated by the type 2A-related Ser/Thr protein phosphatase, Sit4p. The KlPdr5 transporter is a protein of 1,525 amino acids sharing 63.8% sequence identity with its Saccharomyces cerevisiae counterpart, ScPdr5p. Overexpression of the KlPDR5 gene confers resistance to oligomycin, antimycin, econazole, and ketoconazole, whereas cells with a disrupted allele of KlPDR5 are hypersensitive to the drugs and have a decreased capacity to carry out efflux of the anionic fluorescent dye rhodamine 123. It was found that a chromosomal disruption of KlPDR5 abolishes the drug-resistant phenotype associated with sit4 mutations and that a synergistic hyperresistance to the drugs can be created by overexpressing KlPDR5 in sit4 mutants. These data strongly indicate that the multidrug-resistant phenotype of sit4 mutants is mediated by negatively modulating the activity of KlPdr5p. As the transcriptional level of KlPDR5 and the steady-state level of KlPdr5p are not significantly affected by mutations in SIT4, the regulation by Sit4p appears to be a posttranslational process.

ATP-Binding Cassette Transporters↗

PubMed2001

Expression of highly selective sodium channels in alveolar type II cells is determined by culture conditions.

Alveolar fluid clearance in the developing and mature lungs is believed to be mediated by some form of epithelial Na channels (ENaC). However, single-channel studies using isolated alveolar type II (ATII) cells have failed to demonstrate consistently the presence of highly selective Na+ channels that would be expected from ENaC expression. We postulated that in vitro culture conditions might be responsible for alterations in the biophysical properties of Na+ conductances observed in cultured ATII cells. When ATII cells were grown on glass plates submerged in media that lacked steroids, the predominant channel was a 21-pS nonselective cation channel (NSC) with a Na+-to-K+ selectivity of 1; however, when grown on permeable supports in the presence of steroids and air interface, the predominant channel was a low-conductance (6.6 +/- 3.4 pS, n = 94), highly Na+-selective channel (HSC) with a P(Na)/P(K) >80 that is inhibited by submicromolar concentrations of amiloride (K(0.5) = 37 nM) and is similar in biophysical properties to ENaC channels described in other epithelia. To establish the relationship of this HSC channel to the cloned ENaC, we employed antisense oligonucleotide methods to inhibit the individual subunit proteins of ENaC (alpha, beta, and gamma) and used patch-clamp techniques to determine the density of this channel in apical membrane patches of ATII cells. Overnight treatment of cells with antisense oligonucleotides to any of the three subunits of ENaC resulted in a significant decrease in the density of HSC channels in the apical membrane cell-attached patches. Taken together, these results show that when grown on permeable supports in the presence of steroids and air interface, the predominant channels expressed in ATII cells have single-channel characteristics resembling channels that are associated with the coexpression of the three cloned ENaC subunits alpha-, beta-, and gamma-ENaC.

Animals↗

PubMed2001

Pulmonary ventilation imaged by magnetic resonance: at the doorstep of clinical application.

Over the past few years, magnetic resonance imaging (MRI) has emerged as an important instrument for functional ventilation imaging. The aim of this review is to summarize established clinical methods and emerging techniques for research and clinical arenas. Before the advent of MRI, chest radiography and computed tomography (CT) dominated morphological lung imaging, while functional ventilation imaging was accomplished with scintigraphy. Initially, MRI was not used for morphological lung imaging often, due to technical and physical limitations. However, recent developments have considerably improved anatomical MRI, as well as advanced new techniques in functional ventilation imaging, such as inhaled contrast aerosols, oxygen, hyperpolarized noble gases (Helium-3, Xenon-129), and fluorinated gases (sulphur-hexafluoride). Straightforward images demonstrating homogeneity of ventilation and determining ventilated lung volumes can be obtained. Furthermore, new image-derived functional parameters are measurable, such as airspace size, regional oxygen partial pressure, and analysis of ventilation distribution and ventilation/perfusion ratios. There are several advantages to using MRI: lack of radiation, high spatial and temporal resolution and a broad range of functional information. The MRI technique applied in patients with chronic obstructive pulmonary disease, emphysema, cystic fibrosis, asthma, and bronchiolitis obliterans, may yield a higher sensitivity in the detection of ventilation defects than ventilation scintigraphy, CT or standard pulmonary function tests. The next step will be to define the threshold between physiological variation and pathological defects. Using complementary strategies, radiologists will have the tools to characterize the impairment of lung function and to improve specificity.

Animals↗

PubMed2001

Effects of decreasing the frequency of ventilator circuit changes to every 7 days on the rate of ventilator-associated pneumonia in a Beijing hospital.

INTRODUCTION: We investigated whether decreasing ventilator circuit changes from every 2 days to every 7 days would impact ventilator-associated pneumonia rates at our institution. METHODS: All mechanically ventilated patients at Peking Union Medical College Hospital were studied over a 21 month period. From March 1998 to February 1999, ventilator circuits were changed every 2 days, and from June through December 1999, ventilator circuits were changed every 7 days. Nosocomial pneumonia was identified using the criteria of the Centers for Disease Control. RESULTS: In the 2-day-change group, there were 2,277 ventilator-patient days and 38 patients developed pneumonia, resulting in a pneumonia rate of 16.7 cases per 1,000 ventilator days. The 7-day-change group accumulated 972 ventilator days and 8 patients contracted pneumonia, resulting in a pneumonia rate of 8.2 cases per 1,000 ventilator days. The pneumonia rate was significantly lower in the 7-day-change group (p = 0.007). To standardize for seasonal variability, we compared results from the same seasonal time frames (June to December 1998 for the 2-day-change group, and June to December 1999 for the 7-day-change group), and obtained similar findings: during those periods, pneumonia rates were 24.2 cases per 1,000 ventilator days for the 2-day-change group and 8.9 cases per 1,000 ventilator days for the 7-day-change group (p = 0.001). CONCLUSIONS: A circuit change interval of 7 days had a lower risk of ventilator-associated pneumonia than a 2-day change interval. Therefore, ventilator circuits can be safely changed every 7 days in our setting.

Cross Infection↗

PubMed2000

Detection of emphysema in rat lungs by using magnetic resonance measurements of 3He diffusion.

Emphysema is a pulmonary disease characterized by alveolar wall destruction, resulting in enlargement of gas exchange spaces without fibrosis. This condition is a part of chronic obstructive pulmonary disease (COPD), which causes 3.5% of deaths worldwide [Anonymous (1990) World Health Stat. Q. Special, 1-51] and contributes greatly to the global burden of disease [Murray, C. J. & Lopez, A. D. (1996) Science 274, 740-743]. Alveolar regeneration has been shown in animal models and could have potential for clinical treatment of early-stage emphysema. However, current techniques for detection of emphysema are not sensitive at the initial stages. Early-stage human panacinar emphysema is modeled in elastase-treated animals. Here, we provide an in vivo imaging method for differentiating normal and emphysematous rat lungs by measuring the apparent diffusion coefficient (ADC) of hyperpolarized (3)He by using magnetic resonance imaging. These data show that the ADC is significantly larger in elastase-treated rats, indicating alveolar expansion. Whereas these rats were clinically asymptomatic, conventional histology confirmed presence of injury. Our results indicate that measurement of the hyperpolarized (3)He ADC can be a valuable research tool and has potential application in the clinical setting.

Animals↗

PubMed2000

Positive and negative control of multidrug resistance by the Sit4 protein phosphatase in Kluyveromyces lactis.

The nuclear gene encoding the Sit4 protein phosphatase was identified in the budding yeast Kluyveromyces lactis. K. lactis cells carrying a disrupted sit4 allele are resistant to oligomycin, antimycin, ketoconazole, and econazole but hypersensitive to paromomycin, sorbic acid, and 4-nitroquinoline-N-oxide (4-NQO). Overexpression of SIT4 leads to an elevation in resistance to paromomycin and to lesser extent tolerance to sorbic acid, but it has no detectable effect on resistance to 4-NQO. These observations suggest that the Sit4 protein phosphatase has a broad role in modulating multidrug resistance in K. lactis. Expression or activity of a membrane transporter specific for paromomycin and the ABC pumps responsible for 4-NQO and sorbic acid would be positively regulated by Sit4p. In contrast, the function of a Pdr5-type transporter responsible for ketoconazole and econazole extrusion, and probably also for efflux of oligomycin and antimycin, is likely to be negatively regulated by the phosphatase. Drug resistance of sit4 mutants was shown to be mediated by ABC transporters as efflux of the anionic fluorescent dye rhodamine 6G, a substrate for the Pdr5-type pump, is markedly increased in sit4 mutants in an energy-dependent and FK506-sensitive manner.

4-Nitroquinoline-1-oxide↗

PubMed2000

The origin of alveolar macrophages in the transplanted lung: a longitudinal microsatellite-based study of donor and recipient DNA.

Transplanted lungs are initially populated by donor pulmonary alveolar macrophages (PAMs). These will form major antigen presenters for the recipient's suppressed immune system. They may be expected to be replaced by recipient major histocompatibility complex-compatible cells, with time. We have isolated CD14+ PAMs from bronchoalveolar lavage specimens for 6 months after transplantation and identified their origin by using microsatellite analysis. This DNA-based technology permits the reliable identification of the origin of cells from different individuals. We show that replacement of donor PAMs occurs with individual dynamics in each case. Recipient PAMs usually appeared within 2 weeks, whereas donor cells could be retained for as long as 6 months. In this limited series, there was no obvious correlation between the dynamics of this process and the occurrence of rejection episodes or infections.

Adult↗

PubMed2000

Mutant residues suppressing rho(0)-lethality in Kluyveromyces lactis occur at contact sites between subunits of F(1)-ATPase.

Characterisation of 35 Kluyveromyces lactis strains lacking mitochondrial DNA has shown that mutations suppressing rho(0)-lethality are limited to the ATP1, 2 and 3 genes coding for the alpha-, beta- and gamma- subunits of mitochondrial F(1)-ATPase. All atp mutations reduce growth on glucose and three alleles, atp1-2, 1-3 and atp3-1, produce a respiratory deficient phenotype that indicates a drop in efficiency of the F(1)F(0)-ATP synthase complex. ATPase activity is needed for suppression as a double mutant containing an atp allele, together with a mutation abolishing catalytic activity, does not suppress rho(0)-lethality. Positioning of the seven amino acids subject to mutation on the bovine F(1)-ATPase structure shows that two residues are found in a membrane proximal region while five amino acids occur at a region suggested to be a molecular bearing. The intriguing juxtaposition of mutable amino acids to other residues subject to change suggests that mutations affect subunit interactions and alter the properties of F(1) in a manner yet to be determined. An explanation for suppressor activity of atp mutations is discussed in the context of a possible role for F(1)-ATPase in the maintenance of mitochondrial inner membrane potential.

Alleles↗

PubMed2000

Hyperpolarized 3He microspheres as a novel vascular signal source for MRI.

Hyperpolarized (HP) 3He can be encapsulated within biologically compatible microspheres while retaining sufficient polarization to be used as a signal source for MRI. Two microsphere sizes were used, with mean diameters of 5.3 +/- 1.3 microm and 10.9 +/- 3.0 microm. These suspensions ranged in concentration from 0.9-7.0% gas by volume. Spectroscopic measurements in phantoms at 2 T yielded 3He relaxation times that varied with gas concentration. At the highest 3He concentration, the spinlattice relaxation time, T1, was 63.8 +/- 9.4 sec, while the transverse magnetization decayed with a time constant of T2* = 11.0 +/- 0.4 msec. In vivo MR images of the pelvic veins in a rat were acquired during intravenous injection of 3He microspheres (SNR approximately equal 15). Advantages such as intravascular confinement, lack of background signal, and limited recirculation indicate quantitative perfusion measurements may be improved using this novel signal source.

Animals↗

PubMed2000

Mixing oxygen with hyperpolarized (3)He for small-animal lung studies.

Hyperpolarized helium (HP (3)He) is useful for direct MR imaging of the gas spaces of small animal lungs. Previously, breaths of 100% HP (3)He were alternated with breaths of air to maximize helium signal in the lungs and to minimize the depolarizing effects of O(2). However, for high-resolution imaging requiring many HP (3)He breaths (hundreds) and for pulmonary disease studies, a method was needed to simultaneously deliver O(2) and HP (3)He with each breath without significant loss of polarization. We modified our existing computer-controlled ventilator by adding a plastic valve, additional relays and a controller. O(2) and HP (3)He are mixed at the beginning of each breath within the body of a breathing valve, which is attached directly to the endotracheal tube. With this mixing method, we found that T(1) relaxation of HP (3)He in the guinea pig lung was about 20 s compared to 30 s with alternate air/HP (3)He breathing. Because imaging times during each breath are short (about 500 ms), the HP (3)He signal loss from O(2) contact is calculated to be less than 5%. We concluded that the advantages of mixing HP (3)He with O(2), such as shorter imaging times (reduced T(1) losses in reservoir) and improved physiologic stability, outweigh the small signal loss from the depolarizing effects of oxygen on HP (3)He.

Animals↗

PubMed2000

Absence of F1-ATPase activity in Kluyveromyces lactis lacking the epsilon subunit.

The mitochondrial F1-ATPase is a multimeric enzyme, comprised of 3alpha, 3beta, gamma, delta and epsilon subunits, that is primarily responsible for the synthesis of ATP in eukaryotic cells. Recent work has shown that the F1 complex of the petite-negative yeast Kluyveromyces lactis, with specific mutations in the alpha, beta or gamma subunits, has a novel function that suppresses lethality caused by loss of mtDNA. Previously, genes for the four largest subunits of K. lactis F1 have been identified and characterised. In this study the gene coding for the epsilon-subunit of F1, KlATPepsilon, has been isolated and found to encode a polypeptide of 61 amino acids with only 32 residues identical to those in the protein from Sacharomyces cerevisiae. Strains carrying a null mutation of KlATPepsilon are respiratory deficient while the introduction of ATPepsilon from S. cerevisiae restores growth on non-fermentable carbon sources. In contrast to S. cerevisiae, K. lactis disrupted in ATPepsilon does not have a detectable F1-related mitochondrial ATP hydrolysis activity, suggesting that the epsilon-subunit plays a critical role in the formation of the catalytic sector of F1. With a disrupted KlATPepsilon, the rho degrees-lethality suppressor function of F1 carrying the atp2-1 and atp1-6 alleles is abolished. However, inactivation of the epsilon subunit does not eliminate the rho degrees-viable phenotype of the atp1-1, atp2-9, atp3-2 mutants. It is suggested that the absence of epsilon may effect the assembly or stability of F in the wild-type, atp 2-1 and atp1-6 strains, whereas the defect can be suppressed by the atp1-1, atp2-9 and atp3-2 mutations in the alpha, beta and gamma subunits respectively.

Amino Acid Sequence↗

PubMed2000

Disruption of the MRP-L23 gene encoding the mitochondrial ribosomal protein L23 is lethal for Kluyveromyces lactis but not for Saccharomyces cerevisiae.

The Kluyveromyces lactis nuclear gene, MRP-L23, encodes a polypeptide of 155 amino acids that shares 70% and 43% identity to the ribosomal proteins L23 and L13 of Saccharomyces cerevisiae and Escherichia coli. The deduced protein, designated K1L23, is a likely component of the large subunit of mitochondrial ribosomes as it can complement the respiratory deficient phenotype of a S. cerevisiae mrp-L23 mutant. As in S. cerevisiae, KlMRP-L23 is essential for respiratory growth of K. lactis because disruption of the gene in a "petite-positive" strain carrying a rho o-lethality suppressor atp mutation rendered cells unable to grow on a nonfermentable carbon source. However, in contrast to S. cerevisiae, disruption of MRP-L23 in wild type K. lactis is lethal. Meiotic segregants of K. lactis with a disrupted MRP-L23 allele form microcolonies with cell numbers varying from 32 to 300. These data clearly indicate an essential role of mitochondrial protein synthesis for viability of the petite-negative yeast K. lactis.

Amino Acid Sequence↗

PubMed2000

The petite mutation in yeasts: 50 years on.

Fifty years ago it was reported that baker's yeast, Saccharomyces cerevisiae, can form "petite colonie" mutants when treated with the DNA-targeting drug acriflavin. To mark the jubilee of studies on cytoplasmic inheritance, a review of the early work will be presented together with some observations on current developments. The primary emphasis is to address the questions of how loss of mtDNA leads to lethality (rho 0-lethality) in petite-negative yeasts and how S. cerevisiae tolerates elimination of mtDNA. Recent investigation have revealed that rho 0-lethality can be suppressed by specific mutations in the alpha, beta, and gamma subunits of the mitochondrial F1-ATPase of the petite-negative yeast Kluyveromyces lactis and by the nuclear ptp alleles in Schizosaccharomyces pombe. In contrast, inactivation of genes coding for F1-ATPase alpha and beta subunits and disruption of AAC2, PGS1/PEL1, and YME1 genes in S. cerevisiae convert this petite-positive yeast into a petite-negative form. Studies on nuclear genes affecting dependence on mtDNA have provided important insight into the functions provided by the mitochondrial genome and the maintenance of structural and functional integrity of the mitochondrial inner membrane.

Amino Acid Sequence↗