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F Yuan

Publications and source records attributed to F Yuan.

At least 55 records · Page 3Linked to original sources
PubMed1997

Direct in vivo measurement of targeted binding in a human tumor xenograft.

Binding is crucial to the function of most biologically active molecules, but difficult to quantify directly in living tissue. To this end, fluorescence recovery after photobleaching was used to detect the immobilization of fluorescently labeled ligand caused by binding to receptors in vivo. Measurements of mAb affinity to target antigen within human tumor xenografts revealed a saturable binding isotherm, from which an in vivo carcinoembryonic antigen density of 0.56 nmol/g (5.0 x 10(5)/cell) and an association constant of Ka < or = 4 x 10(7) M(-1) were estimated. The present method can be adapted for in vivo studies of cell signaling, targeted drugs, gene therapy, and other processes involving receptor-ligand binding.

Animals↗
PubMed1997

Interlaboratory variation in oxygen tension measurement by Eppendorf "Histograph" and comparison with hypoxic marker.

BACKGROUND AND OBJECTIVES: The median of pO2 values in tumor measured by Eppendorf "Histograph" with a needle-type electrode has been used as a prognostic indicator in cancer patients. However, it is not established that a pretreatment measured pO2 value can be used as a universal predictor of local control probability, because the variation in pO2 values, especially in hypoxic tissue, among institutes may not allow comparison of measured "absolute pO2 values." The purpose of this study was to examine the variation in oxygen tension measurement by Eppendorf "Histograph" among six laboratories using a single batch of mice and tumors and the same detailed protocol. These results were also compared to the immunohistochemical staining of 2-nitromidazole adducts. METHODS: C3H mice bearing FSaII murine fibrosarcoma subcutaneously were shipped to all laboratories, and the oxygen status in tumors and in normal subcutis was examined using Eppendorf "Histograph" and immunohistochemical hypoxic marker. RESULTS: All laboratories showed that the FSaII tumor was hypoxic with at least 77% of measured points under 10 mmHg in pO2 and with a median pO2 value less than that of normal subcutis. These results were further confirmed immunohistochemically. These findings are interpreted as evidence that the pO2 values measured by Eppendorf "Histograph" can be useful. However, the median values of tumor pO2 varied from 1.5 mmHg to 5.6 mmHg among the laboratories, and pO2 of normal subcutis also varied from 28 mmHg to 38 mmHg. There were also significant differences in hypoxic fraction, defined as the fraction under a given oxygen partial pressure (i.e., under 2.5, 5, or 10 mmHg), among institutes. CONCLUSIONS: Caution needs to be exercised in using the absolute, median, or distribution of pO2 values measured by the Eppendorf "Histograph" to compare the data between laboratories or to predict the radiation response in an individual subject.

Animals↗
PubMed1997

Hypertrophy decreases cardiac KATP channel responsiveness to exogenous and locally generated (glycolytic) ATP.

This study tests the hypothesis that glycolytic regulation of KATP channel activity is altered in myocardial hypertrophy. Left ventricular (LV) subendocardial myocytes were isolated from cats with normal or left ventricular hypertrophied hearts (LVH). Saponin-permeabilized open cell-attached patch configurations of normal and LVH cells were exposed to an exogenous ATP consuming system containing hexokinase and 2-deoxyglucose. Phosphoenol pyruvate (PEP, substrate for the last ATP producing step in glycolysis) was applied extracellularly; ADP was present. In both cell types, KATP channels were activated in the absence of PEP, inhibited when PEP was added and activated again when PEP was removed, indicating the cells retained metabolic integrity and generated ATP in the proximity of their KATP channels. Single channel conductance in the absence of PEP was similar (70 pS, normal; 66 pS, LVH). However, LVH KATP channels showed enhanced activity (P0=0.50+/-0.03); normal (0.41+/-0.03) in PEP absence (P<0. 05). PEP responsiveness was reduced in LVH, with IC50, PEP increased to 23 microM; (11 microM normal). Lactate failed to activate KATP channels in both cell types. The concentration-P0 response curves obtained during exposure of open cells to exogenous ATP also revealed reduced responsiveness to ATP of LVH KATP channels (IC50, ATP=283 microM LVH; 93 microM normal). Our data indicate myocardial hypertrophy increases the maximal activity of KATP channels in the absence of ATP and reduces their responsiveness to ATP, including locally generated glycolytic ATP. These alterations in metabolic regulation of myocardial electrophysiology may contribute to diversity of action potential shortening in hypertrophied hearts during acute ischemia.

Adenosine Triphosphate↗
PubMed1997

Interstitial pH and pO2 gradients in solid tumors in vivo: high-resolution measurements reveal a lack of correlation.

The partial pressure of oxygen (pO2) and pH play critical roles in tumor biology and therapy. We report here the first combined, high-resolution (< or = 10 microns) measurements of interstitial pH and pO2 profiles between adjacent vessels in a human tumor xenograft, using fluorescence ratio imaging and phosphorescence quenching microscopy. We found (1) heterogeneity in shapes of pH and pO2 profiles; (2) a discordant relation between local pH profiles and corresponding pO2 profiles, yet a strong correlation between mean pH and pO2 profiles; (3) no correlation between perivascular pH/pO2 and nearest vessel blood flow; and (4) well-perfused tumor vessels that were hypoxic and, consequently, large hypoxic areas in the surrounding interstitium. Such multiparameter measurements of the in vivo microenvironment provide unique insights into biological processes in tumors and their response to treatment.

Adenocarcinoma↗
PubMed1997

Identification of a cytotoxic T-lymphocyte epitope in the human papillomavirus type 16 E2 protein.

Persistent infection with oncogenic types of human papillomaviruses (HPV) is the major cause of cervical cancer precursor lesions. Cellular immune responses are considered important in the elimination of HPV infection, but the targets are not well defined. HPV E1 and E2 proteins form a replicative complex necessary for viral genome maintenance. To investigate whether epitopes in the E1 or E2 proteins can serve as targets for cytotoxic T-lymphocyte (CTL)-mediated killing, we identified peptides containing the human leukocyte antigen (HLA)-A*0201 binding motif in the deduced amino acid sequences of the HPV-16 E1 and E2 genes. Binding affinity of the peptides was measured by HLA-A*0201 up-regulation on T2 cells. Peptides with high binding-affinity were tested for their ability to elicit peptide-specific CTLs from healthy blood donors. We found one peptide from the E1 and one from the E2 protein sequence that were capable of eliciting peptide-specific CTLs. The E2-specific CTLs lysed an HPV-16-transfected cervical carcinoma cell line, but not the untransfected HPV-negative parental cell line, indicating that the identified E2 epitope can be presented to CTLs in HPV-positive epithelial cells. These findings might have potentially important implications for studies of the natural history of HPV infection in relation to cervical carcinogenesis.

Amino Acid Sequence↗
PubMed1997

Regional gradation of L-type calcium currents in the feline heart with a healed myocardial infarct.

INTRODUCTION: Abnormal action potentials in myocytes adjacent to > 2-month-old feline LV myocardial infarcts (MI) may reflect alterations in Ca2+ currents (Ica). METHODS AND RESULTS: We compared ICa, at 36 degrees C, in subendocardial myocytes isolated from areas adjacent to MI and to ICa in cells from remote areas (> 4 mm away; REM) and control cells from similar regions in normal hearts. Control (CON) myocytes had membrane capacitance of 234 +/- 10 pF (n = 81 cells) compared to 305 +/- 14 pF in REM (71 cells; P < 0.05 from CON) and 237 +/- 11 pF (n = 55 cells) in MI (not different from CON). From Vh = -40 mV; peak ICa elicited by test potentials (-35 to +70 mV) were significantly larger in CON (-1746 +/- 123 pA) and REM (-1795 +/- 142 pA) compared to MI (-1352 +/- 129 pA) (P < 0.05). Peak ICa density was significantly reduced in REM (-6.0 +/- 0.4 pA/pF) or MI (-5.7 +/- 0.4 pA/pF, P < 0.05) compared to CON (-7.5 +/- 0.4 pA/pF). Double exponential ICa decay was similar among groups. Half-inactivation potential (V0.5) was significantly shifted (hyperpolarizing direction) for MI (-29.1 +/- 2.6 mV) and REM (-24.6 +/- 1.2 mV) myocytes compared to -20.3 +/- 1.0 mV in CON. MI slope factor (k; 9.0 +/- 0.5) was significantly different from CON (6.8 +/- 0.3) and REM (7.3 +/- 0.4). No differences in time course of recovery from inactivation were noted. Five millimolar Ba2+o produced significant increases in ICa in CON and REM but an attenuated response in MI. Bay K8644 (1 microM) produced similar ICa increase in all groups. ICa increase due to isoproterenol (1 microM) in MI and REM was half that in CON, but there were no differences in increased ICa responses among groups following phenylephrine (10 microM). CONCLUSION: Reduced ICa density in REM reflects cell hypertrophy, whereas altered ICa of MI may reflect altered channel structure and/or function.

Action Potentials↗
PubMed1997

Role of nitric oxide in tumor microcirculation. Blood flow, vascular permeability, and leukocyte-endothelial interactions.

The present study was designed to define the role of nitric oxide (NO) in tumor microcirculation, through the direct intravital microcirculatory observations after administration of NO synthase (NOS) inhibitor and NO donor both regionally and systemically. More specifically, we tested the following hypotheses: 1) endogenous NO derived from tumor vascular endothelium and/or tumor cells increases and/or maintains tumor blood flow, decreases leukocyte-endothelial interactions, and increases vascular permeability, 2) exogenous NO can increase tumor blood flow via vessel dilatation and decrease leukocyte-endothelial interactions, and 3) NO production and tissue responses to NO are tumor dependent. To this end, a murine mammary adenocarcinoma (MCaIV) and a human colon adenocarcinoma (LS174T) were implanted in the dorsal skinfold chamber in C3H and severe combined immunodeficient mice, respectively, and observed by means of intravital fluorescence microscopy. Both regional and systemic inhibition of endogenous NO by N omega-nitro-L-arginine methyl ester (L-NAME; 100 mumol/L superfusion or 10 mg/kg intravenously) significantly decreased vessel diameter and local blood flow rate. The diameter change was dominant on the arteriolar side. Superfusion of NO donor (spermine NO, 100 mumol/L) increased tumor vessel diameter and flow rate, whereas systemic injection of spermine NO (2.62 mg/kg) had no significant effect on these parameters. Rolling and stable adhesion of leukocytes were significantly increased by intravenous injection of L-NAME. In untreated animals, both MCaIV and LS174T tumor vessels were leaky to albumin. Systemic NO inhibition significantly attenuated tumor vascular permeability of MCaIV but not of LS174T tumor. Immunohistochemical studies, using polyclonal antibodies to endothelial NOS and inducible NOS, revealed a diffuse pattern of positive labeling in both MCaIV and LS174T tumors. Nitrite and nitrate levels in tumor interstitial fluid of MCaIV but not of LS174T were significantly higher than that in normal subcutaneous interstitial fluid. These results support our hypotheses regarding the microcirculatory response to NO in tumors. Modulation of NO level in tumors is a potential strategy for altering tumor hemodynamics and thus improving oxygen, drug, gene vector, and effector cell delivery to solid tumors.

Adenocarcinoma↗
PubMed1997

Effect of host microenvironment on the microcirculation of human colon adenocarcinoma.

It is generally accepted that the host microenvironment influences tumor biology. There are discrepancies in growth rate, metastatic potential, and efficacy of systemic treatment between ectopic and orthotopic tumors. Liver is the most common and critical site of distant metastasis of colorectal carcinoma. Tumorigenicity and efficacy of chemotherapeutic agents in colorectal tumors are different in liver and subcutaneous sites. Thus, we hypothesize that the liver (orthotopic) versus subcutaneous (ectopic) microenvironment would have different effects on the angiogenesis and maintenance of the microcirculation of colorectal tumor. To this end, we developed a new method to monitor and to quantify microcirculatory parameters in the tumor grown in the liver. Using this approach, we compared the microcirculation of LS174T, a human colon adenocarcinoma, metastasized to the liver with that of the host liver vessels and that of the same tumor grown in the subcutaneous space. In the liver metastasis model, 5 x 10(6) LS174T cells were injected into the spleen of nude mice. Four to eight weeks later, the liver with metastatic tumors was exteriorized and placed on a special stage and observed under an intravital fluorescence microscope. The dorsal skinfold chamber model was used to study the subcutaneous tumors. Red blood cell velocity, vessel diameter, density, permeability, and leukocyte-endothelial interactions were measured using fluorescence microscopy and image analysis. Vascular endothelial growth factor/ vascular permeability factor (VEGF/VPF) mRNA expression was determined by the Northern blot analysis. LS174T tumor foci in the liver had tortuous vascular architecture, heterogeneous blood flow, significantly lower vascular density, and significantly higher vascular permeability than normal liver tissue. Tumors grown in the liver had significantly lower vessel density, especially in the center coincident with central necrosis, than the subcutaneous tumors. The frequency distribution of vessel diameters of liver tumor was slightly shifted to smaller size compared with that of subcutaneous tumor. Leukocyte rolling in liver tumor was twofold lower than that in subcutaneous tumor. These physiological findings were consistent with the measurement of VEGF/VPF in that the VEGF/VPF mRNA level was lower in the liver tumor than that in the subcutaneous tumor. However, macromolecular vascular permeability in the liver tumor was significantly higher than in the subcutaneous tumor. Liver sinusoidal endothelial cells, the origin of liver tumor vessel endothelium, are known to be fenestrated and not to have a basement membrane, suggesting that the difference in endothelial cell origin may explain the difference in tumor vascular permeability in two sites. These findings demonstrate that liver microenvironment has different effects on some aspects of the tumor angiogenesis and microcirculation compared with the subcutaneous tissues. The new model/method described in this paper has significant implications in two research areas: 1) the liver microenvironment and its effect on tumor pathophysiology in conjunction with cytokine/ growth factor regulation and 2) the delivery of drugs, cells, and genes to liver tumors.

Adenocarcinoma↗
PubMed1996

Time-dependent vascular regression and permeability changes in established human tumor xenografts induced by an anti-vascular endothelial growth factor/vascular permeability factor antibody.

The hyperpermeability of tumor vessels to macromolecules, compared with normal vessels, is presumably due to vascular endothelial growth factor/vascular permeability factor (VEGF/VPF) released by neoplastic and/or host cells. In addition, VEGF/VPF is a potent angiogenic factor. Removal of this growth factor may reduce the permeability and inhibit tumor angiogenesis. To test these hypotheses, we transplanted a human glioblastoma (U87), a human colon adenocarcinoma (LS174T), and a human melanoma (P-MEL) into two locations in immunodeficient mice: the cranial window and the dorsal skinfold chamber. The mice bearing vascularized tumors were treated with a bolus (0.2 ml) of either a neutralizing antibody (A4.6.1) (492 micrograms/ml) against VEGF/VPF or PBS (control). We found that tumor vascular permeability to albumin in antibody-treated groups was lower than in the matched controls and that the effect of the antibody was time-dependent and influenced by the mode of injection. Tumor vascular permeability did not respond to i.p. injection of the antibody until 4 days posttreatment. However, the permeability was reduced within 6 h after i.v. injection of the same amount of antibody. In addition to the reduction in vascular permeability, the tumor vessels became smaller in diameter and less tortuous after antibody injections and eventually disappeared from the surface after four consecutive treatments in U87 tumors. These results demonstrate that tumor vascular permeability can be reduced by neutralization of endogenous VEGF/ VPF and suggest that angiogenesis and the maintenance of integrity of tumor vessels require the presence of VEGF/VPF in the tissue microenvironment. The latter finding reveals a new mechanism of tumor vessel regression-i.e., blocking the interactions between VEFG/VPF and endothelial cells or inhibiting VEGF/VPF synthesis in solid tumors causes dramatic reduction in vessel diameter, which may block the passage of blood elements and thus lead to vascular regression.

Animals↗
PubMed1996

Backbone dynamics, amide hydrogen exchange, and resonance assignments of the DNA methylphosphotriester repair domain of Escherichia coli Ada using NMR.

The 10kDa amino-terminal fragment of Escherichia coli Ada protein (N-Ada10) repairs methyl phosphotriesters in DNA and possesses a tightly bound zinc ion. The complete resonance assignments of this protein domain have been obtained using multidimensional homonuclear and heteronuclear NMR experiments. The assignments served to study the internal mobility of this protein domain via 15N relaxation experiments. This involved the measurement of longitudinal and transverse 15N relaxation rates, as well as the amide proton solvent exchange rates. Relaxation rates in the rotating frame, R1 rho, of 15N nuclei were measured at different spin-lock field strengths, leading to the detection of two slow conformational exchange processes at Gly-25 and Gln-73. For the latter, which is next to the active site of this protein domain, the characteristic time of this process was found to be around 60 microseconds. The other relaxation experiments unveiled some regions of fast internal motions, faster than the overall correlation time. These motions were found in the N- and C- terminal tails, in segment 33-35 which forms the turn between beta-strands S1 and S2, and residues 47-52 located in a long loop preceding strand S3. The latter loop belongs to the potential DNA binding surface of N-Ada10. While the structure from residue 18 to residue 26 appears not well defined in the calculated structure, the relaxation experiments do not indicate higher mobility for this region. Residues at the N-terminal portion, including the first helix, the sequentially adjacent loop, and part of the second helix, exhibit internal motions close to the time scale of the overall rotational correlation time. This appears to be related to the fact that the first helix has no hydrogen bonds or salt bridges to the rest of the protein and is stabilized only by the involvement of some of its side chains in a hydrophobic core consisting of the side chains of two phenylalanines, a tryptophan, a leucine, and a valine. The four cysteines which bind the zinc show motions on different time scales ranging from microseconds to picoseconds. Thus the motions in the immediate region around the bound zinc of the DNA methyl phosphotriester repair domain are of relatively small amplitude but take place over a wide time range. On the other hand, high mobility is found in the turn connecting S1 and S2 and in the loop preceding S3, regions of the potential DNA binding surface.

Bacterial Proteins↗
PubMed1996

Potential Jupiter astmospheric constituents: candidates for the mass spectrometer in the Galileo atmospheric probe.

An array of carbon-hydrogen-nitrogen (CHN) compounds are formed when mixtures of ammonia and acetylene are photolyzed with ultraviolet (UV) light. Some of these compounds, formed in our investigation of pathways for HCN synthesis on Jupiter, have not been encountered previously in observational, theoretical or laboratory photochemical studies. Knowledge of these structures may facilitate the interpretation of the mass spectral data obtained by the Galileo probe as it passed through the atmosphere of Jupiter.

Acetylene↗
PubMed1996

Fluorescence ratio imaging of interstitial pH in solid tumours: effect of glucose on spatial and temporal gradients.

Tumour pH plays a significant role in cancer treatment. However, because of the limitations of the current measurement techniques, spatially and temporally resolved pH data, obtained non-invasively in solid tumours, are not available. Fluorescence ratio imaging microscopy (FRIM) has been used previously for noninvasive, dynamic evaluation of pH in neoplastic tissue in vivo (Martin GR, Jain RK 1994, Cancer Res., 54, 5670-5674). However, owing to problems associated with quantitative fluorescence in thick biological tissues, these studies were limited to thin (50 microns) tumours. We, therefore, adapted the FRIM technique for pH determination in thick (approximately 2 mm) solid tumours in vivo using a pinhole illumination-optical sectioning (PIOS) method. Results show that (1) steep interstitial pH gradients (5 microns resolution), with different spatial patterns, exist between tumour blood vessels; (2) pH decreased by an average of 0.10 pH units over a distance of 40 microns away from the blood vessel wall, and by 0.33 pH units over a 70 microns distance; (3) the maximum pH drop, defined as the pH difference between the intervessel midpoint and the vessel wall, was positively correlated with the intervessel distance; (4) 45 min following a systemic glucose injection (6 g kg-1 i.v), interstitial pH gradients were shifted to lower pH values by an average of 0.15 pH units, while the spatial gradient (slope) was maintained, when compared with preglucose values. This pH decrease was not accompanied by significant changes in local blood flow. pH gradients returned to near-baseline values 90 min after glucose injection; (5) interstitial tumour pH before hyperglycaemia and the glucose-induced pH drop strongly depended on the local vessel density; and (6) sodium bicarbonate treatment, either acute (1 M, 0.119 ml h-1 for 3 h i.v.) or chronic (1% in drinking water for 8 days), did not significantly change interstitial tumour pH. Modified FRIM may be combined with other optical methods (e.g. phosphorescence quenching) to evaluate non-invasively the spatial and temporal characteristics of extracellular pH, intracellular pH and pO2 in solid tumours. This will offer unique information about tumour metabolism and its modification by treatment modalities used in different cancer therapies.

Adenocarcinoma↗
PubMed1996

C/EBPbeta is a negative regulator of human papillomavirus type 11 in keratinocytes.

We have evaluated the impact of the CCAAT enhancer-binding protein (C/EBP) transcription factors on human papillomavirus type 11 (HPV11). C/EBPbeta is in nuclei of cultured foreskin keratinocytes and binds its consensus sequence in HPV11 DNA. We have used the novel approach of depleting the availability of C/EBPs in vivo using nuclease-resistant oligomers containing C/EBP DNA binding sites. In cultured foreskin keratinocytes containing replicating HPV11 DNA, levels of both HPV11 transcripts and HPV DNA increase after treatment with oligomers; containing the C/EBPbeta DNA binding motif. These results indicate that C/EBPbeta is a repressor for HPV11 in keratinocytes.

Base Sequence↗
PubMed1996

Perfusion of single tumor microvessels: application to vascular permeability measurement.

OBJECTIVE: To develop a new method for determining the relative importance of convection versus diffusion in macromolecular transport across tumor microvessel walls. METHODS: The human colon adenocarcinoma LS174T was transplanted in the dorsal skinfold chamber in a severe combined immunodeficient (SCID) mouse. The vasculature at the tumor surface was exposed by carefully removing the glass window of the chamber. A tumor microvessel was randomly selected, which was approximately 20-40 microns in diameter, embedded in the connective tissue 10-12 microns below the surface of the tumor. The vessel was cannulated with a micropipette and perfused with fluorescein isothiocyanate (FITC)-labeled bovine serum albumin (BSA) at different perfusion pressures. The fluorescence intensity was recorded on videotapes via a video system attached to the fluorescence microscope for offline analysis. The apparent vascular permeability was determined based on the time-dependence of fluorescence intensity and the vessel diameter. RESULTS: The apparent vascular permeability of single vessels to FITC-labeled BSA was quantified at perfusion pressures of 20-45 cmH2O. The pressure dependence of vascular permeability in LS174T tumors was heterogeneous. On average, there was no correlation between the apparent vascular permeability and the perfusion pressure in the range of 20-35 cmH2O (p = 0.73), even though the apparent permeability increased significantly when the pressure was increased from 20 to 45 cmH2O (p = 0.008). CONCLUSIONS: These results indicate that convection in the transvascular transport of albumin is not significant in non-peripheral regions of solid tumors in which the pressure difference across the vessel wall is small or even negligible. In addition to the permeability studies, this preparation can be used to study cell-cell interactions in single tumor vessels under defined flow conditions.

Adenocarcinoma↗
PubMed1996

Heating or freezing bone. Effects on angiogenesis induction and growth potential in mice.

We have characterized the effect of bone graft treatment by heating or freezing (with or without dimethyl sulfoxide (DMSO)). Tissue culture and dorsal skin-fold chambers in mice were used as sites to quantify the effect on angiogenesis, growth and calcification of neonatal femora. Fresh femora increased in both length and cartilage diameter (calcification in vivo only), but cryopreservation or heating abolished the increase in femoral dimensions. In vivo, femora of all experimental groups elicited an angiogenic response from the host tissue, which was most pronounced for fresh femora, weaker for DMSO-preserved frozen bone and poor for unprotected frozen bone and boiled femora. Freezing in the presence of a cryopreservative (DMSO) was found to preserve the angiogenic potential of frozen bone, whereas unprotected heating or freezing significantly impaired angiogenesis induction and growth potential.

Animals↗