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Biomedical subjects

Hua Lu

Publications and source records attributed to Hua Lu.

54 records · Page 3Linked to original sources
PubMed2004

Effects of zi chong granules on estradiol synthesis in the cultured ovarian granular cells of mice.

OBJECTIVE: To investigate the effect of Zi Chong Granules ([symbol: see text]) on estradiol (E2) synthesis in the cultured ovarian granular cells of the mice, so as to probe into the mechanism of the drug in promoting follicular development. METHODS: Testosterone (T) with different concentration were added to the cultured ovarian granular cells, and 24 and 48 hours later the enzyme immunoassay (EIA) was used to determine the required concentration of T and time for E2 synthesis; Zi Chong Granules, its active components for tonifying the kidney and removing blood stasis, and the mouse serum containing Zi Chong Granules were respectively added to the medium of the cultured mouse ovarian granular cells and the mixtures were cultured for 48 hours to determine the E2 concentration in various cultures, with insulin growth factor-1 (IGF-1), follicle-stimulating hormone (FSH) and Nü Jin Dan ([symbol: see text] Golden Pill for Women) as positive control drugs. RESULTS: Zi Chong Granules and Nü Jin Dan (were found to be able to increase the E2 level in the medium of the cultured ovarian granular cells to a certain extent, and the active components for tonifying the kidney or those for removing the blood stasis did not increase the E2 level, but the mouse serum containing Zi Chong Granules could significantly increase the level of E2. CONCLUSION: To promote the E2 synthesis in the ovarian granular cells is one of mechanisms of Zi Chong Granules in stimulating follicular development.

Animals↗
PubMed2003

MDM2 promotes p21waf1/cip1 proteasomal turnover independently of ubiquitylation.

The CDK inhibitor p21waf1/cip1 is degraded by a ubiquitin-independent proteolytic pathway. Here, we show that MDM2 mediates this degradation process. Overexpression of wild-type or ring finger-deleted, but not nuclear localization signal (NLS)-deleted, MDM2 decreased p21waf1/cip1 levels without ubiquitylating this protein and affecting its mRNA level in p53(-/-) cells. This decrease was reversed by the proteasome inhibitors MG132 and lactacystin, by p19(arf), and by small interfering RNA (siRNA) against MDM2. p21waf1/cip1 bound to MDM2 in vitro and in cells. The p21waf1/cip1-binding-defective mutant of MDM2 was unable to degrade p21waf1/cip1. MDM2 shortened the half-life of both exogenous and endogenous p21waf1/cip1 by 50% and led to the degradation of its lysine-free mutant. Consequently, MDM2 suppressed p21waf1/cip1-induced cell growth arrest of human p53(-/-) and p53(-/-)/Rb(-/-)cells. These results demonstrate that MDM2 directly inhibits p21waf1/cip1 function by reducing p21waf1/cip1 stability in a ubiquitin-independent fashion.

Adenocarcinoma↗
PubMed2003

Ubiquitination regulates PSD-95 degradation and AMPA receptor surface expression.

PSD-95 is a major scaffolding protein of the postsynaptic density, tethering NMDA- and AMPA-type glutamate receptors to signaling proteins and the neuronal cytoskeleton. Here we show that PSD-95 is regulated by the ubiquitin-proteasome pathway. PSD-95 interacts with and is ubiquitinated by the E3 ligase Mdm2. In response to NMDA receptor activation, PSD-95 is ubiquitinated and rapidly removed from synaptic sites by proteasome-dependent degradation. Mutations that block PSD-95 ubiquitination prevent NMDA-induced AMPA receptor endocytosis. Likewise, proteasome inhibitors prevent NMDA-induced AMPA receptor internalization and synaptically induced long-term depression. This is consistent with the notion that PSD-95 levels are an important determinant of AMPA receptor number at the synapse. These data suggest that ubiquitination of PSD-95 through an Mdm2-mediated pathway is critical in regulating AMPA receptor surface expression during synaptic plasticity.

Acetylcysteine↗
PubMed2003

Ceramides modulate programmed cell death in plants.

The balance between the bioactive sphingolipid ceramide and its phosphorylated derivative has been proposed to modulate the amount of programmed cell death (PCD) in eukaryotes. We characterized the first ceramide kinase (CERK) mutant in any organism. The Arabidopsis CERK mutant, called accelerated cell death 5, accumulates CERK substrates and shows enhanced disease symptoms during pathogen attack and apoptotic-like cell death dependent on defense signaling late in development. ACD5 protein shows high specificity for ceramides in vitro. Strikingly, C2 ceramide induces, whereas its phosphorylated derivative partially blocks, plant PCD, supporting a role for ceramide phosphorylation in modulating cell death in plants.

Apoptosis↗
PubMed2003

Extended pi-conjugated dendrimers based on truxene.

The largest pi-conjugated dendrimers containing up to nine 10,15-dihydro-5H-diindeno[1,2-alpha;1',2';-c]fluorene (truxene) moieties have been prepared with good yields by repetitive Friedel-Crafts acetylation and acid-promoted cyclotrimerization reactions. An alternative approach to the convergent synthesis of desired dendrimers has been developed, in which the core is generated "in-situ" by acid-promoted cyclotrimerizations of aryl methyl ketones. This proves valuable to afford large-size and precisely well-defined dendrimers in an accelerated dendrimer-growth strategy utilizing enlarged repeat units. The increasing amount of SiCl4 dramatically improves the yield of cyclotrimerization reactions. The introduction of hexahexyl groups onto C-5, C-10, C-15 positions of the truxene moiety greatly enhances the solubility of our compounds. Further investigation indicates that the torsion angle between the truxene segment and the benzene ring might play a key role in determining the photo properties of pi-conjugated dendrimers.

Journal Article↗
PubMed2003

Inhibition of endocytosis causes phosphorylation (S256)-independent plasma membrane accumulation of AQP2.

Inhibition of clathrin-mediated endocytosis by expression of a GTPase-deficient dynamin mutant (dynamin-2/K44A) for 16 h results in an accumulation of plasma membrane aquaporin-2 (AQP2) in epithelial cells stably transfected with wild-type AQP2. We now show a similar effect of K44A dynamin in LLC-PK1 cells transfected with an S256 phosphorylation-deficient AQP2 mutant, AQP2(S256A), and in AQP2-transfected inner medullary collecting duct (IMCD) cells. More acute blockade of endocytosis in these cells with the cholesterol-depleting agent methyl-beta-cyclodextrin (mbetaCD; 10 mM) resulted in a rapid and extensive cell-surface accumulation of both wild-type AQP2 and AQP2 (S256A) within 15 min after treatment. This effect was similar to that induced by treatment of the cells with vasopressin. Blockade of endocytosis by mbetaCD was confirmed using quantitative analysis of FITC-dextran uptake and AQP2 membrane insertion was verified by cell-surface biotinylation. These data indicate that AQP2 recycles constitutively and rapidly between intracellular stores and the cell surface in LLC-PK1 and IMCD cells. The constitutive trafficking process is not dependent on phosphorylation of the serine-256 residue of AQP2, which is, however, an essential step for regulated vasopressin/cAMP-mediated translocation of AQP2. Our data show that rapid and extensive plasma membrane accumulation of AQP2 can occur in a vasopressin receptor (V2R)- and phosphorylation-independent manner, pointing to a potential means of bypassing the mutated V2R in X-linked nephrogenic diabetes insipidus to achieve cell surface expression of AQP2.

Adenoviridae↗
PubMed2003

ACD6, a novel ankyrin protein, is a regulator and an effector of salicylic acid signaling in the Arabidopsis defense response.

The previously reported Arabidopsis dominant gain-of-function mutant accelerated cell death6-1 (acd6-1) shows spontaneous cell death and increased disease resistance. acd6-1 also confers increased responsiveness to the major defense signal salicylic acid (SA). To further explore the role of ACD6 in the defense response, we cloned and characterized the gene. ACD6 encodes a novel protein with putative ankyrin and transmembrane regions. It is a member of one of the largest uncharacterized gene families in higher plants. Steady state basal expression of ACD6 mRNA required light, SA, and an intact SA signaling pathway. Additionally, ACD6 mRNA levels were increased in the systemic, uninfected tissue of Pseudomonas syringae-infected plants as well as in plants treated with the SA agonist benzothiazole (BTH). A newly isolated ACD6 loss-of-function mutant was less responsive to BTH and upon P. syringae infection had reduced SA levels and increased susceptibility. Conversely, plants overexpressing ACD6 showed modestly increased SA levels, increased resistance to P. syringae, and BTH-inducible and/or a low level of spontaneous cell death. Thus, ACD6 is a necessary and dose-dependent activator of the defense response against virulent bacteria and can activate SA-dependent cell death.

Ankyrins↗
PubMed2003

Interaction of p53 with cellular proteins.

Cellular proteins that interact with p53 play a major role in both positive and negative regulation of this tumor suppressor and can fine-tune its response to specific cellular stresses. As a consequence, p53 biology will not be complete until these interacting proteins are fully characterized. This chapter outlines two methods for identifying and characterizing p53-binding proteins: (i) glutathione-S-transferase (GST) protein-protein interaction assay; and (ii) co-immunoprecipitation (co-IP) assay. These two methods are ideal for any laboratory to perform; the assays are short in duration, do not require specialized expertise to establish in the laboratory, give reliable and reproducible data, and are cost-efficient, because few reagents are needed. This chapter gives a basic description of these two techniques and provides tips that are not found in other protocol manuals on how to achieve the best results.

Animals↗
PubMed2003

[Human bone marrow mesenchymal stem cells express multiple hematopoietic growth factors].

To study the biological role of human cultured bone marrow mesenchymal stem cell (BM-MSC) in hematopoiesis by investigation of its expression of multiple hematopoietic growth factors, RT-PCR was used to analyze the expression of SCF, Flt3-ligand, TPO, LIF, G-CSF, GM-CSF, IL-3, IL-6 and IL-11 at mRNA level for human BM-MSC from healthy donors and patients with leukemia and lymphoma. BM-MSC were incubated with or without hydrocortison (HC). The results clearly showed that the cultured BM-MSC expressed mRNA of SCF, Flt3-ligand, TPO, LIF, IL-6 and IL-11 at passages 3 up to 15, but did not express G-CSF, GM-CSF and IL-3. The same expression pattern of above cytokines was seen also for the patient's BM-MSC. HC was able to induce BM-MSC to express G-CSF but not to express GM-CSF. BM-MSC seemed not to change morphologically after incubation with HC for up to 21 days. In conclusion, both normal and patient BM-MSC should be potential to promote hematopoiesis according to their expression of multiple hematopoietic cytokines, and HC is able to induce hematopoietic growth factor expression.

Bone Marrow Cells↗
PubMed2002

The acetylase activity of p300 is dispensable for MDM2 stabilization.

It has been shown that p300 binds to MDM2 and leads to down-regulation of the p53 function. However, it remains unclear whether the acetylase activity of p300 is necessary for regulating MDM2 stability. In this study, we address this issue. First, p300 did not acetylate MDM2 in solution and in cells. Second, overexpression of p300 in cells increased the level of the MDM2 protein but not its mRNA. Similarly, the acetylase-defective p300 AT2 mutant stabilized the MDM2 protein as well. Consistently, the deacetylase inhibitor, trichostatin A, did not significantly affect the half-life of the endogenous MDM2 protein, whereas p300 enhanced the half-life of MDM2. Finally, both wild type and acetylase-defective mutant p300 proteins associated with MDM2 in nuclear body-like structures where MDM2 might be protected from proteasomal degradation. Thus, these results suggest that p300 appears to stabilize MDM2 by retaining this protein in a specific nuclear structure rather than by acetylating it.

Acetylation↗
PubMed2002

p53 serine 392 phosphorylation increases after UV through induction of the assembly of the CK2.hSPT16.SSRP1 complex.

Previously, we purified a UV-responsive p53 serine 392 kinase from F9 and HeLa cells and found that its activity is attributed to a high molecular weight protein complex containing the protein kinase CK2, along with the chromatin-associated factors hSPT16 and SSRP1. Here we determine that these proteins interact in vitro and in cells via non-overlapping domains and provide evidence consistent with the idea that hSPT16 and SSRP1 change the conformation of CK2 upon binding such that it specifically targets p53 over other substrates. Also, UV irradiation apparently induces the association of the complex, thereby increasing the specificity of CK2 for p53 at the expense of other cellular CK2 substrates and leading to an overall increase in p53 serine 392 phosphorylation.

Animals↗
PubMed2002

SSRP1 functions as a co-activator of the transcriptional activator p63.

The p53 homolog p63 is a transcriptional activator. Here, we describe the identification of an HMG1-like protein SSRP1 as a co-activator of p63. Over expression of wild-type, but not deletion mutant, SSRP1 remarkably enhanced p63gamma-dependent luciferase activity, G1 arrest, apoptosis and expression of endogenous PIG3, p21(Waf1/cip1) and MDM2 in human p53-deficient lung carcinoma H1299 cells and mouse embryonic fibroblasts. Also, SSRP1 interacted to p63gamma in vitro and in cells, and resided with p63gamma at the p53-responsive DNA element sites of the cellular endogenous MDM2 and p21(Waf1/cip1) promoters. Moreover, N-terminus-deleted p63 (DeltaN-p63) bound to neither SSRP1 nor its central domain in vitro. Accordingly, SSRP1 was unable to stimulate DeltaN-p63-mediated residual luciferase activity and apoptosis in cells. Finally, the ectopic expression of the central p63-binding domain of SSRP1 inhibited p63-dependent transcription in cells. Thus, these results suggest that SSRP1 stimulates p63 activity by associating with this activator at the promoter.

Animals↗
PubMed2002

MDM2 inhibits PCAF (p300/CREB-binding protein-associated factor)-mediated p53 acetylation.

Our previous study shows that MDM2, a negative feedback regulator of the tumor suppressor p53, inhibits p300-mediated p53 acetylation. Because PCAF (p300/CREB-binding protein-associated factor) also acetylates and activates p53 after DNA damage, in this study we have examined the effect of MDM2 on PCAF-mediated p53 acetylation. We have found that MDM2 inhibited p53 acetylation by PCAF in vitro. In addition, when overexpressed, MDM2 inhibited PCAF-mediated p53 acetylation in cells. MDM2 interacted with PCAF both in vitro and in cells, as assessed using GST fusion protein interaction and immunoprecipitation assays, respectively. Consistent with the above results, MDM2 significantly repressed the activation of p53 transcriptional activity by PCAF without apparently affecting the level of p53. In addition, MDM2 co-resided with p53 at the p53-responsive mdm2 and p21(waf1/cip1) promoters, inhibiting expression of the endogenous p21(waf1/cip1). These results demonstrate that MDM2 can inhibit PCAF-mediated p53 acetylation and activation.

Acetylation↗
PubMed2002

[Inhibitory effect of all-trans-retinoid and polyphenon-100 on microsatellite instability in a colon cancer line].

OBJECTIVE: To investigate the effects of all-trans-retinoic acid(ATRA) and polyphenon-100 (PP) on genetic instability of human tumor cells via their role in alteration of microsatellite sequence(MS) and the expression of mismatch repair gene hMLH(1) and hMSH(2) in RER(+) (replication error) cells. METHODS: RER(+) colon cancer cell line was used as a host for lipofection with pCMV-CAR in which a foreign (CA)(14) repeat was inserted in the coding sequence of LacZ reporter gene, resulting in misreading LacZ frame. Any mutation which made the base number of (CA)(14) tract to be 3-fold resumed normal reading frame of LacZ, and thus led to expression of beta-galactosidase. Variable expression of LacZ in the transfectant cells resulting from RATA or PP treatment was measured by OD reading at lambda 620 after X-gal staining. Expression of mismatch repair genes of hMLH(1) and hMSH(2) was examined at mRNA level by reverse transcription-polymerase chain reaction (RT-PCR). RESULTS: ATRA at 1 mu mol/L, 0.1 u mol/L and PP at 3 mu g/ml had no significant inhibitory effect on cell proliferation. After being treated with ATRA or PP for 1 week, the blue cells of RKO transfectant clones were significantly reduced, and this meant the mutation of exogenous (CA)(14) in RKO cells were inhibited. But no expression of hMLH(1) and hMSH(2) was observed. CONCLUSION: The above data showed both ATRA and PP had inhibitory effects on MS instability of cancer and thus demonstrated directly their beneficial role in stabilization of genomic DNA. However, the present authors have not observed any expression of hMLH(1) and hMSH(2) in RKO cells treated with ATRA or PP.

Adaptor Proteins, Signal Transducing↗
PubMed2002

Transplantation of amniotic epithelial cells into fetal rat liver by in utero manipulation.

It has been hoped that amniotic epithelial cells would be a gene carrier to neural and hepatic tissue, because of 1) the presence of neural and hepatic stem-like cells, 2) the ability to cryopreserve them, 3) long-term survival in the transplanted site, and 4) few ethical problems concerning procurement. But transplantation of a sufficient number of cells to adult tissue needs large-scale cell supply and may lead to vascular embolism. We attempted transplantation of amniotic epithelial cells into fetal liver, because 1) the fetal liver is at the proliferative stage, 2) the number of cells required is small, and 3) the fetal stage is advantageous for the induction of immunological tolerance. Amniotic epithelial cells from day 18.5-20.5 fetuses were transfected with adenoviral AdlacZ and harvested to inject into fetal rat liver of the syngeneic strain (day 18.5-20.5). The efficacy of cell transplantation into the liver increased in the order: intraplacental < intraumbilical vein < intrahepatic route. LacZ-transfected amniotic cells (1-8 x 10(5) cells), hepatocytes (5 x 10(5) cells), or AdlacZ vector solution (1.7 x 10(7) pfu) were injected through the uterine membrane into the liver. Transplanted cells formed a cellular mass and survived for up to 14 days after birth, whereas lacZ-transfected cells were rapidly decreased after the injection of AdlacZ vector or rat hepatocytes as a gene carrier so that the use of amniotic epithelial cells as a gene carrier will result in long-term expression of exogenous genes in the liver.

Adenoviridae↗
PubMed2002

[Clinicopathologic features and diagnosis of metastatic carcinoma to the spleen].

OBJECTIVE: To study the clinicopathologic features and diagnosis of metastatic carcinoma to the spleen (MCS). METHODS: Four patients (1 man and 3 women, mean age 43.5 years) with MCS were analyzed clinicopathologically. RESULTS: The four MCS patients accounted for 1.3% of 308 patients having spleen biopsy from 1959 to 1999. Their chief presentations were pain and mass in the left upper quadrant of the abdomen. The mass was located in the upper pole of the spleen (1 patient), the lower pole of the spleen (2), or the lower pole and hilum of the spleen (1). Macroscopically, all of the lesions were nodular. Histologic type of these MCSs included acinar cell carcinoma of the pancreas (2 patients), transverse colon adenocarcinoma (1), and hepatic cell carcinoma (1). Clinically, 1 patient was diagnosed as having MCS and 3 were misdiagnosed. According to Chinese literature, the clinicopathologic features of MCS were as follows: (1) 66.7% of the patients with MCS were aged 30 approximately 60 years, with a mean of 51.2 years. (2) 76.3% of the patients presented with pain in the left upper quadrant of the abdomen and 63.2% with splenomegaly and splenic masses. (3) Macroscopically, nodular lesions accounted for 68.4%. (4) Microscopically, 84.2% of the lesions were adenocarcinomas and 70.3% originated from carcinomas of the colon, liver, ovary and pancreas. (5) B-mode ultrasonography and/or CT showed occupying lesions or masses in the spleen in 76.7%, and MCS in 11.8%. (6) Clinically, 73.7% of the patients were misdiagnosed. CONCLUSIONS: MCS is uncommon but its clinical misdiagnosis rate is high. Image examination is of value in clinical diagnosis. Cooperation of clinicians and pathologists may enhance the diagnostic level of MCS.

Adult↗
PubMed2001

Developmental expression and perturbation of arabinogalactan-proteins during seed germination and seedling growth in tomato.

Arabinogalactan-proteins (AGPs) are a family of highly glycosylated hydroxyproline-rich glycoproteins present throughout the plant kingdom. A synthetic chemical reagent, (beta-D-Gal)3 Yariv reagent, specifically binds AGPs and can be used for histochemical staining, isolating and probing the function of AGPs. Here, the role of AGPs in tomato (Lycopersicon esculentum Mill. cv. UC82B) seed germination and seedling growth was examined by following expression of AGPs during these events and by treatment with (beta-D-Gal)3 Yariv to perturb AGP function. AGP expression changed during germination and seedling development both quantitatively and qualitatively as revealed by analysis of total AGP content, crossed electrophoresis patterns, RNA blots using LeAGP-1 probe, and western blots with LeAGP-1, JIM13, and MAC207 antibodies. (beta-D-Gal)3 Yariv treatment of seeds and developing seedlings did not affect percent seed germination, but markedly inhibited seedling growth in roots and to a lesser degree in shoots. Root growth inhibition encompassed reductions in overall root length, epidermal root cell elongation, root cell numbers and root hair formation. This growth inhibition was reversible following removal of (beta-D-Gal)3 Yariv. In a related experiment, water uptake by tomato seedlings was greatly inhibited by (beta-D-Gal)3 Yariv treatment. Based on these experiments, AGPs are clearly associated with tomato seedling development and likely to function in root growth, more specifically in cell elongation, cell proliferation, root hair formation and water uptake.

Journal Article↗