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Biomedical subjects

J Lu

Publications and source records attributed to J Lu.

At least 163 records · Page 9Linked to original sources
PubMed2001

[Optimization of the preparation of 3',5'-dioctanoyl-5-fluoro-2'-deoxyuridine pharmacosomes using central composite design].

AIM: To optimize the preparation of 3', 5'-dioctanoyl-5-fluoro-2'-deoxyuridine pharmacosomes (DO-FUdR-PS) by using central composite design. METHODS: DO-FUdR-PS was prepared by a thin-layer ultrasonication technique. The effects of drug phosphatidycholine ratio, pluronic F-68 concentration (%, w/v) and glycerol tristearate (%, w/v) concentration on the mean particle size, entrapment ratio (ER) and drug loading (DL) were investigated. A second-order polynomial equation was fitted to the data and the resulting model was used to predict the response in the optimal region. RESULTS: All the investigated response variables were found to be highly dependent on the formulation variables. Under the optimized conditions, the mean particle size, ER and DL of DO-FUdR-PS were 76 nm, 97.49% and 31.44%, respectively, which highly agreed with the predicted values. CONCLUSION: Central composite design was successfully used to optimize the preparation of DO-FUdR-PS.

Drug Design↗
PubMed2001

[Effect of Rg2 on hemodynamics of hemorrhagic shock and its antioxidation in dogs].

OBJECTIVE: To investigate the protective effect of Rg2 on hemodynamics of hemorrhagic shock and its antioxidant properties. METHOD: Twenty mongrel dogs were randomly divided into Rg2 group, Shen Mai group and control group. The hemorrhagic shock model was built in all dogs by artery bleeding and mean arterial blood pressure was kept < 5.33 kPa for 4.5 hours. Rg2 0.5-1.0 mg.kg-1 and Shen Mai 100 mg.kg-1 were intravenously administered after hemorrhagic shock appeared. RESULT: Rg2 significantly enhanced blood pressure, LVSP and +/- dp/dtmax on hemorrhagic shock dogs who had lost compensation ability. Rg2 0.5-1.0 mg.kg-1 could reduce serum MDA, increase superoxide dismutase activity and prolong survival rate of dogs after shock. All these data have statistically significant when compared to control group (P < 0.05, P < 0.01, P < 0.001). CONCLUSION: These results suggest that Rg2 is more potent and effective than Shen Mai in improving hemodynamic state and activiting SOD on hemorrhagic shock dogs.

Animals↗
PubMed2001

[Regulatory effects of mifepristone and progesterone on the secretion of interleukin-6 by cultured eutopic and etopic endometrial cells].

OBJECTIVE: To investigate the regulatory effects of mifepristone and progesterone on the secretion of interleukin-6 (IL-6) by endometrial and endometriosis cells in vitro. METHODS: Primary cultures of eutopic and ectopic endometrial cells from 9 cases of endometriosis were exposed to mifepristone (10(-6) mol/L, 10(-4) mol/L) and progesterone (1 x 10(-7) mol/L, 1 x 10(-5) mol/L) respectively. IL-6 secretion was analyzed in the culture medium by enzyme linked immunosorbent assay (ELSA). RESULTS: Mifepristone inhibited the IL-6 secretion of ectopic endometrial cells, with the concentrations of IL-6 was (1 914.33 +/- 799.28) microg/L in the 1 x 10(-6) mol/L group (P < 0.01) and (990.25 +/- 58.40) microg/L in the 10(-4) mol/L group (P < 0.01). Progesterone also inhibited IL-6 secretion of ectopic endometrial cells, with (2 575.89 +/- 119.75) microg/L in the 1 x 10(-7) mol/L group (P < 0.05) and (1 736.25 +/- 750.89) microg/L in the 10(-5) mol/L group (P < 0.01). Mifepristone restrained the secretion of IL-6 by eutopic endometrial cells, with (346.96 < or = 24.32) microg/L in the 1 x 10(-6) mol/L group (P < 0.01) and (270.22 +/- 36.15) microg/L in the 1 x 10(-4) mol/L group (P < 0.01). All the effects of down regulation were more obvious when the concentrations of mifepristone and progesterone increased (P < 0.05 - 0.01) . Secretion of IL-6 by eutopic endometrial cells did not change significantly when incubated with progesterone (P > 0.05). CONCLUSION: The inhibitory effects on the secretion of IL-6 by ectopic and (or) eutopic endometrium may provide one of the cellular therapeutic mechanisms of mifepristone and progesterone on endometriosis.

Cells, Cultured↗
PubMed2001

[Mifepristone following conservative surgery in the treatment of endometriosis].

OBJECTIVE: To compare the efficacy and safety of mifepristone and danazol after conservative surgery in the treatment of patients with endometriosis. METHODS: Sixty-one patients with endometriosis (RAFS stage I-IV ) after conservative surgery were treated orally either with mifepristone 10 mg/d (group M, n = 31) or danazol 200 mg 2-3 times/d (group D, n = 30) for 3 months. Changes of symptoms and signs, serum reproductive hormone levels as well as side effects were assessed before and at the end of therapy. Moreover, biochemical parameters of bone metabolism: urinary deoxypyridine /creatinine (UDpd/Cr), serum alkaline phosphatase (AKP) and bone gala-protein (BGP) were also measured before and after treatments. RESULTS: During treatment symptoms and signs were remarkbly relieved in both groups. Side effects including hot flushes, irregular vaginal bleeding, back pain, weight gain and acne, were less commnly seen in group M as compared with group D. Serum luteal hormone (LH), follicular stimulating homone (FSH) levels remained in the range of follicular phase in both groups. So was serum estradiol (E2) levels in group M[(204.9 +/- 45.3 ) pmol/L], but declined to postmenopausal level in group D [(94.3 +/- 33.0) pmol/L]. About two weeks after discontinuation of the thrapy, serum E2 levels [(1,221.6 +/- 384.2) pmol/L] was not significantly different from the normal ovulatory range in group M, but significantly lower in group D [(815.1 +/- 376.0) pmo/L, P < 0.05] . So were the serum progesterone levels at mid-luteal phase [(33.1 +/- 5.6) nmol/L Vs (27.4 +/- 4.9) nmol/L]. There were no significant changes of biochemical parameters of bone metabolism before and at the end of treatment except significant increases of serum AKP and BGP in group D. CONCLUSIONS: Mifepristone is equally effective to danazol when combined with conservative surgery in the management of endometriosis with fewer side effects. Serum E2 levels remained in the range of follicular phase. No impact on the bone turnover after 3 months of therapy was found.

Bone Remodeling↗
PubMed2000

Ligand- and coactivator-mediated transactivation function (AF2) of the androgen receptor ligand-binding domain is inhibited by the cognate hinge region.

Transactivation functions (AF2) in the ligand-binding domains (LBD) of many steroid receptors are well characterized, but there is little evidence to support such a function for the LBD of the androgen receptor (AR). We report a mutant AR, with residues 628-646 in the hinge region deleted, which exhibited transactivation activity that was more than double that of the wild type (WT) AR. Although no androgen-dependent AF2 activity could be observed for the WT ARLBD fused to a heterologous DNA-binding domain, the mutant ARLBD(Delta628-646) was 30-40 times more active than the WT ARLBD. In the presence of the p160 coactivator TIF2, AR(Delta628-646) was significantly more active than similarly treated WT AR. Deletion of residues 628-646 also enhanced TIF2-ARLBD activity 8-fold, an effect not present when the LBD-interacting LXXLL motifs of TIF2 were mutated, suggesting that the negative modulatory activity of residues 628-646 were exerted via coactivator pathways. Although the AP-1 (c-Jun/c-Fos) system and NcoR have been reported to interact with and repress the activity of some steroid receptors, c-Jun, c-Fos, c-Jun/c-Fos, nor NcoR function was consistently affected by the absence or presence of residues 628-646, implying that the AR hinge region exerts its silencing effects in a manner independent of these corepressors. Our data provide evidence for the novel finding that strong androgen-dependent AF2 exists in the ARLBD and is the first report of a negative regulatory domain in the AR. Because mutations in this region are commonly associated with prostate cancer, it is important to characterize the mechanisms by which the hinge region exerts its repressor effect on ligand-activated and coactivator-mediated AF2 activity of the ARLBD.

Amino Acid Sequence↗
PubMed2000

Assessment of the sensitivity and specificity of oligonucleotide (50mer) microarrays.

To examine the utility and performance of 50mer oligonucleotide (oligonucleotide probe) microarrays, gene-specific oligonucleotide probes were spotted along with PCR probes onto glass microarrays and the performance of each probe type was evaluated. The specificity of oligonucleotide probes was studied using target RNAs that shared various degrees of sequence similarity. Sensitivity was defined as the ability to detect a 3-fold change in mRNA. No significant difference in sensitivity between oligonucleotide probes and PCR probes was observed and both had a minimum reproducible detection limit of approximately 10 mRNA copies/cell. Specificity studies showed that for a given oligonucleotide probe any 'non-target' transcripts (cDNAs) >75% similar over the 50 base target may show cross-hybridization. Thus non-target sequences which have >75-80% sequence similarity with target sequences (within the oligonucleotide probe 50 base target region) will contribute to the overall signal intensity. In addition, if the 50 base target region is marginally similar, it must not include a stretch of complementary sequence >15 contiguous bases. Therefore, knowledge about the target sequence, as well as its similarity to other mRNAs in the target tissue or RNA sample, is required to design successful oligonucleotide probes for quality microarray results. Together these results validate the utility of oligonucleotide probe (50mer) glass microarrays.

Animals↗
PubMed2000

Differential effects of theaflavin monogallates on cell growth, apoptosis, and Cox-2 gene expression in cancerous versus normal cells.

Theaflavin (TF-1), theaflavin-3-monogallate and theaflavin-3'-monogallate mixture (TF-2), and theaflavin-3,3'-digallate (TF-3) are the major black tea polyphenols. Here we compared the effects of these polyphenols on cell growth, apoptosis, and gene expression in normal and cancerous cells. We showed that TF-2 (10-50 microM) inhibited the growth of SV40 transformed WI38 human cells (WI38VA) and Caco-2 colon cancer cells but had little effect on the growth of their normal counterparts. The IC50s of TF-2 for the growth inhibition of WI38 and WI38VA cells were, respectively, 300 and 3 microM. The other two black tea polyphenols, TF-1 and TF-3, did not exhibit such differential growth-inhibitory effect. TF-2, but not TF-1 or TF-3, induced apoptosis in transformed WI38VA cells but not in normal WI38 cells, suggesting that apoptosis was responsible, at least in part, for the differential growth-inhibitory effect of TF-2. Cox-2 has been implicated in intestinal carcinogenesis. Among the tea polyphenols tested, TF-2 and, to a lesser degree, (-)-epigallocatechin gallate inhibited cyclooxygenase (Cox)-2 gene expression. TF-2 at 50 microM completely blocked the serum-induced Cox-2 gene expression at both mRNA and protein level. Other genes, including c-fos, c-myc, thymidine kinase, proliferating cell nuclear antigen, BRCA1, BRCA2, and Cox-1, were not significantly affected by TF-2. These findings suggest that TF-2 may be responsible, at least in part, for the chemopreventive activity in black tea extracts.

Antioxidants↗
PubMed2000

Aminohexanoic hydroxamate is a potent inducer of the differentiation of mouse neuroblastoma cells.

Deoxyhypusine synthase is the key enzyme for modifying a lysine residue to hypusine in the cellular protein eukaryotic initiation factor 5A (eIF-5A). Deletion of the deoxyhypusine synthase or the eIF-5A gene in yeast produces lethal phenotype. Inhibition of deoxyhypusine synthase by 1-guanidino-7-aminoheptane (GC7) suppresses tumor cell growth. Hypusine formation represents one of the most specific polyamine-dependent biochemical reactions. In view of the importance of polyamines in growth regulation and cancer biology, deoxyhypusine synthase has been considered to be a good target for chemotherapeutic drug design. Using GC7 as a prototype we have synthesized and tested three classes of diamine analogs, namely, guanidino-, pyrimidino-, and hydroxamate derivatives, as potential inhibitors for deoxyhypusine synthase. Our study shows that (i) among all the compounds tested, GC7 remained to be the most potent inhibitor for deoxyhypusine synthase; (ii) N,N'-bispyrimidino-1, 9-diaminononane, although a poor inhibitor of deoxyhypusine synthase, was a potent growth inhibitor; and (iii) one of the hydroxamate derivatives, 6-aminohexanoic hydroxamate (HC6), prominently induced the differentiation of mouse neuroblastoma cells at sub-millimolar concentrations. Interestingly, other hydroxamates with different chain length were not nearly as effective as HC6 in inducing neuroblastoma cell differentiation. The effect of HC6 was also unique in that it could induce neurite outgrowth and the expression of neuron-specific genes such as synapsin I and MAP-2 in neuroblastoma cells in the absence of other promoting agents such as cAMP. The effect of HC6 on neuroblastoma cell differentiation was comparable with, or better than that of N(6),O(2)'-dibutyryl cAMP (Bt(2)cAMP), a standard reagent commonly used for inducing the differentiation of mouse and human neuroblastoma cells in culture.

3T3 Cells↗
PubMed2000

Signal-dependent nuclear export of a histone deacetylase regulates muscle differentiation.

Members of the myocyte enhancer factor-2 (MEF2) family of transcription factors associate with myogenic basic helix-loop-helix transcription factors such as MyoD to activate skeletal myogenesis. MEF2 proteins also interact with the class II histone deacetylases HDAC4 and HDAC5, resulting in repression of MEF2-dependent genes. Execution of the muscle differentiation program requires release of MEF2 from repression by HDACs, which are expressed constitutively in myoblasts and myotubes. Here we show that HDAC5 shuttles from the nucleus to the cytoplasm when myoblasts are triggered to differentiate. Calcium/calmodulin-dependent protein kinase (CaMK) signalling, which stimulates myogenesis and prevents formation of MEF2-HDAC complexes, also induces nuclear export of HDAC4 and HDAC5 by phosphorylation of these transcriptional repressors. An HDAC5 mutant lacking two CaMK phosphorylation sites is resistant to CaMK-mediated nuclear export and acts as a dominant inhibitor of skeletal myogenesis, whereas a cytoplasmic HDAC5 mutant is unable to block efficiently the muscle differentiation program. Our results highlight a mechanism for transcriptional regulation through signal- and differentiation-dependent nuclear export of a chromatin-remodelling enzyme, and suggest that nucleo-cytoplasmic trafficking of HDACs is involved in the control of cellular differentiation.

Animals↗
PubMed2000

Pseudosubstrate inhibition of protein kinase PKR by swine pox virus C8L gene product.

The interferon-induced protein kinase PKR is activated upon binding double-stranded RNA and phosphorylates the translation initiation factor eIF2alpha on Ser-51 to inhibit protein synthesis in virally infected cells. Swinepox virus C8L and vaccinia virus K3L gene products structurally resemble the amino-terminal third of eIF2alpha. We demonstrate that the C8L protein, like the K3L protein, can reverse the toxic effects caused by high level expression of human PKR in yeast cells. In addition, expression of either the K3L or C8L gene product was found to reverse the inhibition of reporter gene translation caused by PKR expression in mammalian cells. The inhibitory function of the K3L and C8L gene products in these assays was found to be critically dependent on residues near the carboxyl-termini of the proteins including a sequence motif shared among eIF2alpha and the C8L and K3L gene products. Thus, despite significant sequence differences both the C8L and K3L proteins function as pseudosubstrate inhibitors of PKR.

3T3 Cells↗
PubMed2000

Distinct cAMP response element-binding protein (CREB) domains stimulate different steps in a concerted mechanism of transcription activation.

Hormones and neurotransmitters rapidly change patterns of gene expression in target cells by activating protein kinases that phosphorylate and modify the activity of CREB and other transcription factors. Although CREB was initially characterized as mediating the response to cAMP, CREB phosphorylation and activation are stimulated by diverse extracellular signals and protein kinases in essentially all cells and tissues. CREB stimulates transcription through a constitutive activation domain (CAD), which interacts with the promoter recognition factor TFIID, and through a kinase-inducible domain (KID), when Ser-133 is phosphorylated. The present study provides new insight into the mechanism of activation by showing that each of the CREB domains contributes to transcription initiation by stimulating sequential steps in the transcription reaction. The CAD effectively assembled a polymerase complex, as evidenced by constitutive activation in vivo and stimulation of single-round transcription in vitro. In contrast, phosphorylation of the KID in CREB stimulated isomerization of the polymerase complex, as determined by abortive initiation, and promoter clearance and/or reinitiation, as measured by multiple rounds of transcription. Our results provide evidence for a new model for CREB-mediated induction through a concerted mechanism involving establishment of a polymerase complex by the CAD, followed by stimulation of isomerization, promoter clearance, and/or reinitiation by phosphorylated KID to enhance target gene transcription.

Cell Line↗
PubMed2000

N-NO bond dissociation energies of N-nitroso diphenylamine derivatives (or analogues) and their radical anions: implications for the effect of reductive electron transfer on N-NO bond activation and for the mechanisms of NO transfer to nitranions

The heterolytic and homolytic N-NO bond dissociation energies [i.e., deltaHhet(N-NO) and deltaHhomo(N-NO)] of 12 N-nitroso-diphenylamine derivatives (1-12) and two N-nitrosoindoles (13 and 14) in acetonitrile were determined by titration calorimetry and from a thermodynamic cycle, respectively. Comparison of these two sets of data indicates that homolysis of the N-NO bonds to generate NO* and nitrogen radical is energetically much more favorable (by 23.3-44.8 kcal/mol) than the corresponding heterolysis to generate a pair of ions, giving hints for the driving force and possible mechanism of NO-initiated chemical and biological transformations. The first (N-NO)-* bond dissociation energies [i.e., deltaH(N-NO)-* and deltaH'(N-NO)-*] of radical anions 1-*-14-* were also derived on the basis of appropriate cycles utilizing the experimentally measured deltaHhet(N-NO) and electrochemical data. Comparisons of these two quantities with those of the neutral N-NO bonds indicate a remarkable bond activation upon a possible one-electron transfer to the N-NO bonds, with an average bond-weakening effect of 48.8 +/- 0.3 kcal/mol for heterolysis and 22.3 +/- 0.3 kcal/mol for homolysis, respectively. The good to excellent linear correlations among the energetics of the related heterolytic processes [deltaHhet(N-NO), deltaH(N-NO)-*, and pKa(N-H)] and the related homolytic processes [deltaHhomo(N-NO), deltaH'(N-NO)-*, and BDE(N-H)] imply that the governing structural factors for these bond scissions are similar. Examples illustrating the use of such bond energetic data jointly with relevant redox potentials for analyzing various mechanistic possibilities for nitrosation of nitranions are presented.

Journal Article↗
PubMed2000

Use of two predictive algorithms of the world wide web for the identification of tumor-reactive T-cell epitopes.

Tumor cells can be effectively recognized and eliminated by CTLs. One approach for the development of CTL-based cancer immunotherapy for solid tumors requires the use of the appropriate immunogenic peptide epitopes that are derived from defined tumor-associated antigens. Because CTL peptide epitopes are restricted to specific MHC alleles, to design immune therapies for the general population it is necessary to identify epitopes for the most commonly found human MHC alleles. The identification of such epitopes has been based on MHC-peptide-binding assays that are costly and labor-intensive. We report here the use of two computer-based prediction algorithms, which are readily available in the public domain (Internet), to identify HL4-B7-restricted CTL epitopes for carcinoembryonic antigen (CEA). These algorithms identified three candidate peptides that we studied for their capacity to induce CTL responses in vitro using lymphocytes from HLA-B7+ normal blood donors. The results show that one of these peptides, CEA9(632) (IPQQHTQVL) was efficient in the induction of primary CTL responses when dendritic cells were used as antigen-presenting cells. These CTLs were efficient in killing tumor cells that express HLA-B7 and produce CEA. The identification of this HLA-B7-restricted CTL epitope will be useful for the design of ethnically unbiased, widely applicable immunotherapies for common solid epithelial tumors expressing CEA. Moreover, our strategy of identifying MHC class I-restricted CTL epitopes without the need of peptide/HLA-binding assays provides a convenient and cost-saving alternative approach to previous methods.

Algorithms↗
PubMed2000

Lipopolysaccharide activates specific populations of hypothalamic and brainstem neurons that project to the spinal cord.

Sympathetic preganglionic neurons receive direct, monosynaptic input from a series of well defined nuclei in the brainstem and the hypothalamus. These premotor cell groups coordinate sympathetic control with ongoing endocrine and behavioral response. However, it is not known precisely which populations of sympathetic premotor neurons are activated during specific responses, such as fever after intravenous lipopolysaccharide (LPS). We used the activation of c-fos protein expression in spinally projecting neurons during intravenous LPS fever as a model for examining the functional organization of this system. Intravenous LPS (5 microg/kg) induced Fos-like immunoreactivity in sympathetic preganglionic neurons in the spinal cord as well as several sympathetic premotor nuclei, including the paraventricular nucleus of the hypothalamus, rostral and caudal levels of the ventrolateral medulla, and the nucleus of the solitary tract. After injecting Fluorogold into the intermediolateral column at the T1-L1 spinal levels, neurons that were both Fos immunoreactive and retrogradely labeled were found only in the dorsal parvicellular division of the paraventricular nucleus in the hypothalamus, the rostral ventrolateral medulla (C1 adrenergic cell group), and the A5 noradrenergic cell group in the brainstem. The same pattern of double-labeling was seen from injections at each spinal cord level. These findings suggest that only a limited pool of hypothalamo-sympathetic neurons contribute to the fever response and that they may do so by contacting specific populations of preganglionic neurons that are distributed across a wide range of spinal levels. The anatomical specificity of the paraventriculo-spinal projection is thus functional rather than topographic.

Animals↗
PubMed2000

Changes in apoptosis-related protein (p53, Bax, Bcl-2 and Fos) expression with DNA fragmentation in the central nervous system in rats after closed head injury.

This study aimed to examine the temporal profile of neuronal apoptosis in the central nervous system (CNS) following closed head injury in rat. Fos immunoreactivity was detected in neuronal nuclei in the cerebral cortex at 2 h after head injury. At 4 h, Bax protein expression was elevated with a concomitant down-regulation of Bcl-2 expression. Along with this, a marked immunoexpression of p53 was also observed in these cells. Double immunolabelling study has shown the colocalization of Bax immunoreactivity with Bcl-2 and p53. In rats killed 1 day after injury, a variable number of transferase d-UTP nick-end labelling positive cells were observed. Present findings suggest that the upregulation of p53 and a shift in the ratio of Bcl-2 to Bax may contribute to neuronal apoptosis in the CNS after closed head injury.

Animals↗
PubMed2000

Cytochrome c release and apoptosis induced by mitochondrial targeting of nuclear orphan receptor TR3.

TR3, an immediate-early response gene and an orphan member of the steroid-thyroid hormone-retinoid receptor superfamily of transcription factors, regulates apoptosis through an unknown mechanism. In response to apoptotic stimuli, TR3 translocates from the nucleus to mitochondria to induce cytochrome c release and apoptosis. Mitochondrial targeting of TR3, but not its DNA binding and transactivation, is essential for its proapoptotic effect. Our results reveal a mechanism by which a nuclear transcription factor translocates to mitochondria to initiate apoptosis.

Apoptosis↗
PubMed2000

The basic helix-loop-helix transcription factor capsulin controls spleen organogenesis.

Formation of numerous internal organs involves reciprocal epithelial-mesenchymal signaling and subsequent patterning and growth of the organ primordium. Capsulin is a basic helix-loop-helix transcription factor expressed in mesenchymal cells that encapsulate the epithelial primordia of internal organs, including the kidney and lung, as well as the epicardium, which gives rise to the coronary arteries. Capsulin is also expressed in the mesothelium that gives rise to the spleen. We demonstrate that mice homozygous for a capsulin null mutation fail to form a spleen. The homeobox genes Hox11 and Bapx1, shown previously to be essential regulators of spleen organogenesis, and a lacZ reporter introduced into the capsulin locus, were expressed in the early splenic primordium, derived from the splanchnic mesoderm, of homozygous mutant embryos. However, this primordium failed to develop beyond an initial group of precursor cells and underwent rapid apoptosis. The phenotype of capsulin mutant mice demonstrates that capsulin acts within a subpopulation of splanchnic mesodermal cells to control an essential early step in spleen organogenesis that is likely to represent a point of regulatory convergence of the capsulin, Hox11, and Bapx1 genes.

Animals↗