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Biomedical subjects

Y Guo

Publications and source records attributed to Y Guo.

At least 163 records · Page 9Linked to original sources
PubMed2000

Direct observations in vivo on the role of endothelial selectins and alpha(4) integrin in cytokine-induced leukocyte-endothelium interactions in the mouse aorta.

The molecular mechanisms underlying leukocyte recruitment in large arteries have been extensively studied using histological techniques on fixed tissues. However, there are no reports that address the dynamics of leukocyte recruitment in large arteries in vivo. We developed an intravital microscopy technique for direct observation of leukocyte-endothelium interactions in the mouse aorta. Circulating leukocytes were labeled intravasally with rhodamine 6G and microscopically visualized within the aorta, allowing direct analysis of leukocyte rolling and adhesion. In untreated vessels, leukocyte-endothelium interactions were virtually absent. However, local pretreatment with cytokines interleukin-1beta and tumor necrosis factor-alpha induced clear-cut leukocyte rolling and adhesion, compatible with normal blood flow and wall shear rate. High shear decreased rolling leukocyte flux and increased leukocyte rolling velocity, thus decreasing the tendency for firm adhesion. Leukocyte rolling was almost abolished by an antibody blocking the function of P-selectin, whereas function-blocking antibodies against E-selectin and the alpha(4)-integrin subunit decreased rolling leukocyte flux to 51+/-34% (mean +/- SD) and 59+/-11% of the value before antibody treatment, respectively. In addition, inhibition of E-selectin function, but not of alpha(4) integrin, resulted in increased leukocyte rolling velocity (from 48+/-32 to 71+/-32 microm per second). Taken together, we introduce the first model for direct studies of leukocyte-endothelium interactions in a large artery in vivo and demonstrate cytokine-induced shear-sensitive leukocyte rolling that is critically dependent on P-selectin and modulated by E-selectin and alpha(4) integrin.

Animals↗
PubMed2000

Hybrid cell vaccination for cancer immune therapy: first clinical trial with metastatic melanoma.

Hybrid cell vaccination is a new cancer immune therapy approach that aims at recruiting T cell help for the induction of tumour specific cytolytic immunity. The vaccines are generated by fusion of the patients' tumour cells with allogeneic MHC class II bearing cells to combine the tumour's antigenicity with the immunogenicity of allogeneic MHC molecules. Safety and anti-tumour activity of this treatment were assessed in a clinical trial that has yielded one complete and one partial remission, and 5 cases of stable disease among 16 patients with advanced stage metastatic melanoma. As evidenced by histology, the vaccination induced T cell relocation into tumour nodules. Stable disease could be maintained by repeated booster injections for more than 24 months in some patients. The side effects were minor. Occasional occurrences of vitiligo spots after vaccination were indicative of a restricted therapy induced auto-immune reactivity. The results suggest that hybrid cell vaccination is a safe cancer immune therapy potentially effective for induction of acute anti-tumour response as well as long-term maintenance.

Adult↗
PubMed2000

Racial differences in prostate cancer growth: apoptosis and cell proliferation in Caucasian and African-American patients.

BACKGROUND: Epidemiologic evidence reveals striking racial differences in incidence and clinical behavior of prostate cancer among American men. In this study, we assessed the incidence of apoptosis and cell proliferation in prostate cancer specimens from African-American and Caucasian patients in an attempt to identify potential differences in tumor growth determinants between the two ethnic groups. METHODS: Apoptosis and cell proliferation were analyzed in archival paraffin-embedded prostatic tumors from 44 African-American and 35 Caucasian age-matched men who underwent radical prostatectomy for localized prostate cancer. Both groups had comparable preoperative prostate-specific antigen (PSA) levels, clinical stage, and Gleason scores, and neither group of patients received neoadjuvant therapy prior to surgery. Apoptotic status in prostate tumors was evaluated in situ, using the transferase deoxyuridine end labeling (TUNEL) assay, and the expression profile of two apoptotic proteins, bcl-2 and bax. The proliferative index was determined on the basis of Ki-67 antigen immunoreactivity. RESULTS: Apoptosis in malignant prostate cells was significantly higher in African American than Caucasian men (11.6% vs. 4.2%, P < 0. 001). Interestingly, the rate of cell proliferation of prostate tumor cells was similar in the two ethnic groups (4.5% and 4.2%). The antiapoptotic protein bcl-2 was detected at significantly higher levels in tumors from Caucasian than African-American patients (40. 8% vs. 31.6%, P < 0.05). Expression of bax, the apoptosis promoter, was consistently high among tumor epithelial cells in specimens from both racial groups (68%). CONCLUSIONS: These findings provide a novel insight into the molecular determinants of tumor growth that may underlie the ethnic differences in prostate cancer incidence and clinical behavior. Downregulation of bcl-2 expression may be potentially responsible for the loss of apoptotic control in prostate tumors from African-American men. This study may have significant clinical implications in the development of novel diagnostic approaches for biologically aggressive prostate cancer from diverse racial origin.

Aged↗
PubMed2000

Behavioral responses to cocaine and amphetamine administration in mice lacking the dopamine D1 receptor.

Cocaine and amphetamine can induce both short-term and long-term behavioral changes in rodents. The major target for these psychostimulants is thought to be the brain dopamine system. To determine whether the dopamine D1 receptor plays a crucial role in the behavioral effects of psychostimulants, we tested both the locomotor and stereotyped behaviors in D1 receptor mutant and wild-type control mice after cocaine and amphetamine treatments. We found that the overall locomotor responses of D1 receptor mutant mice to repeated cocaine administration were significantly reduced compared to those of the wild-type mice and the responses of the D1 receptor mutant mice to cocaine injections were never significantly higher than their responses to saline injections. D1 receptor mutant mice were less sensitive than the wild-type mice to acute amphetamine stimulation over a dose range even though they exhibited apparently similar behavioral responses as those of the wild-type mice after repeated amphetamine administration at the 5 mg/kg dose. Immunostaining experiments indicated that there was no detectable neurotoxicity in the nucleus accumbens in both D1 receptor mutant and wild-type mice after repeated amphetamine administration. The data suggest that the D1 receptor plays an essential role in mediating cocaine-induced behavioral changes in mice. Moreover, the D1 receptor also participates in behavioral responses induced by amphetamine administration.

Amphetamine↗
PubMed2000

Physical characteristics and chemical degradation of amorphous quinapril hydrochloride.

This study was designed to investigate the relationships between the solid-state chemical instability and physical characteristics of a model drug, quinapril hydrochloride (QHCl), in the amorphous state. Amorphous QHCl samples were prepared by rapid evaporation from dichloromethane solution and by grinding and subsequent heating of the crystalline form. Physical characteristics, including the glass transition temperature and molecular mobility, were determined using differential scanning calorimetry, thermogravimetric analysis, powder x-ray diffractometry, polarizing microscopy, scanning electron microscopy, and infrared spectroscopy. The amorphous form of QHCl, produced by both methods, has a T(g) of 91 degrees C. Isothermal degradation studies showed that cyclization of QHCl occurred at the same rate for amorphous samples prepared by the two methods. The activation energy was determined to be 30 to 35 kcal/mol. The rate of the reaction was shown to be affected by sample weight, dilution through mixing with another solid, and by altering the pressure above the sample. The temperature dependence for chemical reactivity below T(g) correlated very closely with the temperature dependence of molecular mobility. Above T(g), however, the reaction was considerably slower than predicted from molecular mobility. From an analysis of all data, it appears that agglomeration and sintering of particles caused by softening of the solid, particularly above T(g), and a resulting reduction of the particle surface/volume ratio play a major role in affecting the reaction rate by decreasing the rate of removal of the gaseous HCl product.

Angiotensin-Converting Enzyme Inhibitors↗
PubMed2000

Spontaneous leukocyte rolling in rat and mouse microvessels is independent of mast cell activity.

OBJECTIVE AND DESIGN: The role of mast cells in spontaneous leukocyte rolling in venules of the mouse cremaster muscle and rat mesentery was investigated. MATERIALS: The experiments were carried out using mast cell-deficient rats (Ws/Ws), WBB6F1 mice (W/Wv), and their congenic littermates (wild type). TREATMENT: Administration of compound 48/80 intraperitoneally (50 microg) in rats and intrascrotally (5 microg) in mice, 4 h prior to the experiments. METHODS: Intravital microscopy of the terminal vascular beds in mouse cremaster muscle and rat mesentery. RESULTS: The level of spontaneous leukocyte rolling and the rolling velocity in venules of mast cell-deficient animals exactly matched that seen in wild-type animals. Challenge with compound 48/80 markedly increased leukocyte adhesion and emigration in venules of wild-type animals. In contrast, the number of adherent and extravascular leukocytes was very low in compound 48/80-challenged animals lacking mast cells and did not differ from that seen in control animals treated with phosphate-buffered saline. CONCLUSIONS: The presence or activation of mast cells has no bearing on spontaneous leukocyte rolling, at least not in rat and mouse microvessels.

Animals↗
PubMed2000

Evidence for an essential role of cyclooxygenase-2 as a mediator of the late phase of ischemic preconditioning in mice.

Recent studies have demonstrated that cyclooxygenase-2 (COX-2) is an essential mediator of the cardioprotective effects of the late phase of ischemic preconditioning (PC) in rabbits. The goal of this study was to determine whether COX-2 also plays an essential role in late PC in the mouse. B6129F2/J mice underwent a 30-min coronary occlusion followed by 24 h of reperfusion. Administration of the COX-2 selective inhibitor, NS-398, 30 min prior to the 30-min occlusion (5 mg/kg i.p.) had no appreciable effect on infarct size compared with untreated controls (58.8 +/- 2.1%, vs. 58.8 +/- 4.3% of the risk region, respectively). When mice were preconditioned with six cycles of 4-min coronary occlusion/4-min reperfusion 24 h prior to the 30-min occlusion, infarct size was markedly reduced (19.3 +/- 3.4%), indicating a late PC effect. The protective effect of late PC was completely abrogated by administration of NS-398 30 min before the 30-min coronary occlusion (67.7 +/- 3.0%), but not by administration of vehicle alone (23.6 +/- 3.7%). These results indicate that COX-2 mediates the late phase of ischemic PC in the mouse and imply that the role of this enzyme in cardioprotection is not species-specific.

Animals↗
PubMed2000

SM5-1: a new monoclonal antibody which is highly sensitive and specific for melanocytic lesions.

Antibodies such as HMB-45 and anti-S100 protein have been widely used as markers of malignant melanoma despite evidence that HMB-45 has a sensitivity of only 67-93% and S100 is nonspecific for melanoma. Using a subtractive immunization protocol in a mouse model of human melanoma, we have generated several monoclonal antibodies with putative specificity for melanoma. After initial screenings, the antibody SM5-1 was chosen because of its intriguing reactivity with melanocytic tumors in both frozen and paraffin sections. The immunohistochemical staining of SM5-1 was studied in paraffin-embedded specimens of 401 melanomas (n = 401; 250 primary melanomas, 151 metastases), melanocytic nevi of the skin (n = 16), nonmelanocytic neoplasms (n = 84). The results were compared with HMB-45 and anti-S100 staining. All antibodies reacted with nevi and 97-99% with primary melanomas. Whereas both SM5-1 and anti-S100 stained 96% (146/151) of melanoma metastases, HMB-45 correctly identified only 83% (126/151). All HMB-45-negative metastases were positive for SM5-1. Whereas neither SM5-1 nor HMB-45 stained any of 84 specimens from 40 different nonmelanocytic neoplasms, anti-S100 was positive in 21/84 (25%). While the staining pattern of SM5-1 was mostly homogeneous, small tumor areas in some metastases remained unstained. Staining with SM5-1 was also observed in perivascular dendritic cells, in plasma cells, some myofibroblasts and the secretion of eccrine sweat glands. Nonactivated epidermal melanocytes, keratinocytes, endothelial cells, smooth muscle cells and peripheral nerves were all negative for SM5-1. These results suggest that SM5-1 is highly specific, as well as sensitive, for melanocytic lesions and is useful in the immunohistochemical evaluation of melanoma.

Animals↗
PubMed2000

The Fn14 immediate-early response gene is induced during liver regeneration and highly expressed in both human and murine hepatocellular carcinomas.

Polypeptide growth factors stimulate mammalian cell proliferation by binding to specific cell surface receptors. This interaction triggers numerous biochemical responses including the activation of protein phosphorylation cascades and the enhanced expression of specific genes. We have identified several fibroblast growth factor (FGF)-inducible genes in murine NIH 3T3 cells and recently reported that one of them, the FGF-inducible 14 (Fn14) immediate-early response gene, is predicted to encode a novel, cell surface-localized type Ia transmembrane protein. Here, we report that the human Fn14 homolog is located on chromosome 16p13.3 and encodes a 129-amino acid protein with approximately 82% sequence identity to the murine protein. The human Fn14 gene, like the murine Fn14 gene, is expressed at elevated levels after FGF, calf serum or phorbol ester treatment of fibroblasts in vitro and is expressed at relatively high levels in heart and kidney in vivo. We also report that the human Fn14 gene is expressed at relatively low levels in normal liver tissue but at high levels in liver cancer cell lines and in hepatocellular carcinoma specimens. Furthermore, the murine Fn14 gene is rapidly induced during liver regeneration in vivo and is expressed at high levels in the hepatocellular carcinoma nodules that develop in the c-myc/transforming growth factor-alpha-driven and the hepatitis B virus X protein-driven transgenic mouse models of hepatocarcinogenesis. These results indicate that Fn14 may play a role in hepatocyte growth control and liver neoplasia.

3T3 Cells↗
PubMed2000

Effects of a pectic polysaccharide from a medicinal herb, the roots of Bupleurum falcatum L. on interleukin 6 production of murine B cells and B cell lines.

The effect of T-cell-independent B cell mitogen, a pectic polysaccharide, bupleuran 2IIc, from a medicinal herb, the roots of Bupleurum falcatum L. on interleukin 6 (IL-6) production of murine B cells and B cell lines was investigated in order to clarify the mechanism of enhanced immunoglobulin (Ig) secretion from B cells. Bupleuran 2IIc enhanced IgM secretion from highly purified murine normal B cells. When normal B cells from murine spleen were cultured with bupleuran 2IIc in the presence of anti-IL-6 neutralizing antibody, the enhanced IgM secretion by bupleuran 2IIc was reduced. When B cells were stimulated with bupleuran 2IIc, their IL-6 secretion and the transcription of IL-6 mRNA were enhanced. The enhanced IL-6 secretion by bupleuran 2IIc was also observed in B cell line, Y16 cell. When bupleuran 2IIc was digested with endo-alpha-(1-->4)-D-polygalacturonase, the resulting enzyme resistant carbohydrate portion, "ramified" region (PG-1) composed of rhamnogalacturonan core containing neutral sugar side chains showed potent IL-6 secretion-enhancing activity. These results suggest that the "ramified" region of bupleuran 2IIc stimulates the secretion of IL-6 as the active site, and the resulting IL-6 may partially contribute the enhancement of IgM secretion as an autocrine and/or paracrine mechanism.

Adjuvants, Immunologic↗
PubMed2000

Junction phosphate is derived from the precursor in the tRNA spliced by the archaeon Haloferax volcanii cell extract.

RNA splicing in archaea requires at least an endonuclease and a ligase, as is the case for the splicing of eukaryal nuclear tRNAs. Splicing endonucleases from archaea and eukarya are homologous, although they differ in subunit composition and substrate recognition properties. However, they all produce 2',3' cyclic phosphate and 5'-hydroxyl termini. An in vitro-transcribed, partial intron-deleted Haloferax volcanii elongator tRNA(Met) has been used to study splicing by H. volcanii cell extracts. Substrates and products were analyzed by nearest neighbor analyses using nuclease P1 and RNase T2, and fingerprinting analyses using acid-urea gels in the first dimension and gradient thin layer chromatography in the second dimension. The results suggest that 2',3' cyclic phosphate at the 3' end of the 5' exon is converted into the splice junction phosphate forming a 3',5'-phosphodiester linkage. This resembles the animal cell type systems where the junction phosphate preexists in the transcript, and differs from yeast type systems, where GTP is the source of junction phosphate.

Base Sequence↗
PubMed2000

Effects of lyophilization on the physical characteristics and chemical stability of amorphous quinapril hydrochloride.

PURPOSE: To prepare amorphous quinapril hydrochloride (QHCl) by lyophilization and to compare its physical characteristics and chemical stability as a function of the initial pH of the pre-lyophilized solution. METHODS: Amorphous QHCl samples were prepared by lyophilization from aqueous solutions. Solid-state characteristics were evaluated by DSC, PXRD, and optical microscopy. Chemical degradation was monitored by an HPLC assay. RESULTS: Amorphous QHCl samples obtained from lyophilization exhibited variable glass transition temperatures, depending on the pH and/or concentration of the starting aqueous solutions. Neutralized quinapril (Q) in the amorphous form, which has a Tg of 51 degrees C, lower than that of its HCl salt (91 degrees C), was significantly more reactive than QHCl at the same temperature. The Tg of lyophilized samples prepared at various initial pH values correlated well with values predicted for mixtures of QHCl and Q. Their different reaction rates were related to their glass transition temperature, consistent with the results from earlier studies obtained with amorphous samples made by precipitation from an organic solution and grinding of the crystal solvate. CONCLUSIONS: Lyophilization of different QHCl solutions produces mixtures of amorphous QHCl and its neutralized form Q, with Tg values intermediate to the values of QHCl and Q. As the fraction of Q increases the overall rate of chemical degradation increases relative to QHCl alone, primarily due to the increase in molecular mobility induced by the plasticizing effects of Q.

Algorithms↗
PubMed2000

Enhancement of immunogenicity of tumor cells by cotransfection with genes encoding antisense insulin-like growth factor-1 and B7.1 molecules.

Insulin-like growth factor-1 (IGF-1) is expressed in many tumor cell lines and has a role in both normal cell proliferation and in the growth of cancers. Tumor cells transfected with a vector encoding an IGF-1 antisense cDNA transcriptional cassette driven by the mouse metallothionein-1 promoter become immunogenic and lose their tumorigenicity in syngeneic animals. The enhanced immunogenicity is associated with an up-regulation in the expression of major histocompatibility complex class I molecule on cell surfaces. Blockade of the expression of IGF-1 in tumor cells by the IGF-1 antisense RNA approach is not uniformly effective in the induction of antitumoral protective immunity in low and nonimmunogenic tumor model systems. Here, we report that the immunogenicity of hepa 1-6 hepatoma and SMCC-1 colon carcinoma cells, which are poorly immunogenic and unresponsive to antisense IGF-1 gene transfer, can be induced by cotransfection with genes encoding antisense IGF-1 and mouse B7.1 molecules. The tumor cells modified in this manner become strongly immunogenic and can be used as a cellular vaccine to induce a protective immune response in vivo. Immunization with the transfected tumor cells also results in regression of the established hepa 1-6 hepatoma and SMCC-1 colon cancer. The immunity is tumor-specific and is mediated by CD3+ CD8+ T cells. Cytotoxic T lymphocytes generated in vitro by priming naive spleen cells and in vivo by immunizing mice with the double-transfected tumor cells specifically lysed autologous tumors cells and were effective in adoptive immunotherapy. The data suggest that modification of tumor cells in vitro by cotransfection with genes encoding antisense IGF-1 and B7.1 molecules may open a new avenue for cancer immunogene therapy.

Adjuvants, Immunologic↗
PubMed2000

Lymphocyte function antigen 1 (LFA-1) mediates early tumour necrosis factor alpha-induced leucocyte adhesion in venules.

A co-ordinated expression of specific adhesion molecules regulates leucocyte-endothelium interactions in the microcirculation. In the present study, we used intravital microscopy of the cremaster muscle in CD11a gene-targeted mice to examine the role of lymphocyte function antigen 1 (LFA-1, CD11a/CD18) in tumour necrosis factor alpha (TNF-alpha)-induced leucocyte rolling and firm adhesion in venules. We found that 30 min after TNF-alpha administration leucocyte rolling was unchanged compared with phosphate-buffered saline (PBS)-treated mice and similar in LFA-1-deficient and wild-type animals. In contrast, firm leucocyte adhesion in venules increased by almost 10-fold following 30 min of TNF-alpha challenge in LFA-1-expressing animals, whereas no increase was observed in LFA-1-deficient mice. Four hours after intrascrotal administration of TNF-alpha, venular leucocyte adhesion was found to be markedly increased, but similar in extent to LFA-1-deficient and wild-type mice. Histological examination of haematoxylin-eosin-stained tissue sections revealed that approximately 90% of the leucocytes in the TNF-alpha-stimulated venules in both wild-type and LFA-1-deficient mice were polymorphonuclear. Taken together, our functional in vivo data demonstrate that LFA-1 is an important adhesion molecule in early TNF-alpha-induced venular leucocyte adhesion.

Animals↗
PubMed2000

The N-terminal region of the heme-regulated eIF2alpha kinase is an autonomous heme binding domain.

The N-terminal domain (NTD) of the heme-regulated eukaryotic initiation factor (eIF)2alpha kinase (HRI) was aligned to sequences in the NCBI data base using ENTREZ and a PAM250 matrix. Significant similarity was found between amino acids 11-118 in the NTD of rabbit HRI and amino acids 16-120 in mammalian alpha-globins. Several conserved amino acid residues present in globins are conserved in the NTD of HRI. His83 of HRI was predicted to be equivalent to the proximal heme ligand (HisF8) that is conserved in all globins. Molecular modeling of the NTD indicated that its amino acid sequence was compatible with the globin fold. Recombinant NTD (residues 1-159) was expressed in Escherichia coli. Spectral analysis of affinity purified recombinant NTD indicated that the NTD contained stably bound hemin. Mutational analysis indicated that His83 played a critical structural role in the stable binding of heme to the NTD, and was required to stabilize full length HRI synthesized de novo in the rabbit reticulocyte lysate. These results indicate that the NTD of HRI is an autonomous heme-binding domain, with His83 possibly serving as the proximal heme binding ligand.

Amino Acid Sequence↗
PubMed2000

Control of cellular phosphatidylinositol 4,5-bisphosphate levels by adhesion signals and rho GTPases in NIH 3T3 fibroblasts involvement of both phosphatidylinositol-4-phosphate 5-kinase and phospholipase C.

The involvement of small GTPases of the Rho family in the control of phosphoinositide metabolism by adhesion signals was examined in NIH 3T3 fibroblasts. Abrogation of adhesion signals by detachment of cells from their substratum resulted in a time-dependent decrease in the cellular level of PtdIns(4,5)P2 by approximately 50%. This effect could be mimicked by treatment of adherent cells with Clostridium difficile toxin B and toxin B-1470, which inhibit specific subsets of Rho and Ras GTPases. Detachment of cells that had been pretreated with the clostridial toxins did not cause a further reduction in PtdIns(4,5)P2 levels, suggesting that the target GTPases are integrated into the control of phosphoinositide levels by adhesion signals. The reduction in PtdIns(4,5)P2 levels could be attributed to reduced activity of the major PtdIns(4, 5)P2-producing enzyme, PtdIns4P 5-kinase. Unexpectedly, both cell detachment and toxin treatment resulted in a twofold to threefold increase in inositol phosphate production in intact cells. In lysates of these cells, in vitro phospholipase C activity was found to be elevated by 30-50%. The effects of cell detachment and toxin treatment on inositol phosphate formation could be mimicked by expression of dominant-negative N17 Rac1. Taken together, these data suggest that adhesion-controlled small GTPases of the Rho family are involved in the regulation of the cellular PtdIns(4,5)P2 levels in NIH 3T3 fibroblasts, by controlling the activities of both PtdIns4P 5-kinase and phospholipase C.

3T3 Cells↗