Biomedical subjects

N Wang

Publications and source records attributed to N Wang.

At least 199 records · Page 11Linked to original sources

PubMed1998

Targeted mutation reveals a central role for SR-BI in hepatic selective uptake of high density lipoprotein cholesterol.

Scavenger receptor BI (SR-BI) is a cell surface receptor that binds high density lipoproteins (HDL) and mediates selective uptake of HDL cholesteryl esters (CE) in transfected cells. To address the physiological role of SR-BI in HDL cholesterol homeostasis, mice were generated bearing an SR-BI promoter mutation that resulted in decreased expression of the receptor in homozygous mutant (designated SR-BI att) mice. Hepatic expression of the receptor was reduced by 53% with a corresponding increase in total plasma cholesterol levels of 50-70% in SR-BI att mice, attributable almost exclusively to elevated plasma HDL. In addition to increased HDL-CE, HDL phospholipids and apo A-1 levels were elevated, and there was an increase in HDL particle size in mutant mice. Metabolic studies using HDL bearing nondegradable radiolabels in both the protein and lipid components demonstrated that reducing hepatic SR-BI expression by half was associated with a decrease of 47% in selective uptake of CE by the liver, and a corresponding reduction of 53% in selective removal of HDL-CE from plasma. Taken together, these findings strongly support a pivotal role for hepatic SR-BI expression in regulating plasma HDL levels and indicate that SR-BI is the major molecule mediating selective CE uptake by the liver. The inverse correlation between plasma HDL levels and atherosclerosis further suggests that SR-BI may influence the development of coronary artery disease.

Animals↗

PubMed1998

Differences in elasticity of vinculin-deficient F9 cells measured by magnetometry and atomic force microscopy.

We have investigated a mouse F9 embryonic carcinoma cell line, in which both vinculin genes were inactivated by homologous recombination, that exhibits defective adhesion and spreading [Coll et al. (1995) Proc. Natl. Acad. Sci. USA 92, 9161-9165]. Using a magnetometer and RGD-coated magnetic microbeads, we measured the local effect of loss and replacement of vinculin on mechanical force transfer across integrins. Vinculin-deficient F9Vin(-/-) cells showed a 21% difference in relative stiffness compared to wild-type cells. This was restored to near wild-type levels after transfection and constitutive expression of increasing amounts of vinculin into F9Vin(-/-) cells. In contrast, the transfection of vinculin constructs deficient in amino acids 1-288 (containing the talin- and alpha-actinin-binding site) or substituting tyrosine for phenylalanine (phosphorylation site, amino acid 822) in F9Vin(-/-) cells resulted in partial restoration of stiffness. Using atomic force microscopy to map the relative elasticity of entire F9 cells by 128 x 128 (n = 16,384) force scans, we observed a correlation with magnetometer measurements. These findings suggest that vinculin may promote cell adhesions and spreading by stabilizing focal adhesions and transferring mechanical stresses that drive cytoskeletal remodeling, thereby affecting the elastic properties of the cell.

Animals↗

PubMed1998

Scavenger receptor class B type I as a mediator of cellular cholesterol efflux to lipoproteins and phospholipid acceptors.

We recently reported that the rate of efflux of cholesterol from cells to high density lipoprotein (HDL) was related to the expression level of scavenger receptor class B type I (SR-BI). Moreover, the expression of this receptor in atheromatous arteries raises the possibility that SR-BI mediates cholesterol efflux in the arterial wall (Ji, Y., Jian, B., Wang, N., Sun, Y., de la Llera Moya, M., Phillips, M. C., Rothblat, G. H., Swaney, J. B., and Tall, A. R. (1997) J. Biol. Chem. 272, 20982-20985). In this paper we describe studies that suggest that the presence of phospholipid on acceptor particles plays an important role in modulating interaction with the SR-BI. Specifically, enrichment of serum with phospholipid resulted in marked stimulation of cholesterol efflux from cells that had higher levels of SR-BI expression, like Fu5AH or Y1-BS1 cells, and little or no stimulation in cells with low SR-BI levels, such as Y-1 cells. Stimulation of efflux by phospholipid enrichment was also a function of SR-BI levels in Chinese hamster ovary cells transfected with the SR-BI gene. Efflux to protein-free vesicles prepared with 1-palmitoyl-2-oleoylphosphatidyl-choline also correlated with SR-BI levels, suggesting that phospholipid, as well as protein, influences the interaction that results in cholesterol efflux. By contrast, cholesterol efflux from a non-cell donor showed no stimulation consequent to phospholipid enrichment of the serum acceptor. These results may help to explain observations in the literature that document an increased risk of atherosclerosis in patients with depressed levels of HDL phospholipid even in the face of normal HDL cholesterol levels.

Animals↗

PubMed1998

Phloem Transport of Fructans in the Crassulacean Acid Metabolism Species Agave deserti

Four oligofructans (neokestose, 1-kestose, nystose, and an un-identified pentofructan) occurred in the vascular tissues and phloem sap of mature leaves of Agave deserti. Fructosyltransferases (responsible for fructan biosynthesis) also occurred in the vascular tissues. In contrast, oligofructans and fructosyltransferases were virtually absent from the chlorenchyma, suggesting that fructan biosynthesis was restricted to the vascular tissues. On a molar basis, these oligofructans accounted for 46% of the total soluble sugars in the vascular tissues (sucrose [Suc] for 26%) and for 19% in the phloem sap (fructose for 24% and Suc for 53%). The Suc concentration was 1.8 times higher in the cytosol of the chlorenchyma cells than in the phloem sap; the nystose concentration was 4.9 times higher and that of pentofructan was 3.2 times higher in the vascular tissues than in the phloem sap. To our knowledge, these results provide the first evidence that oligofructans are synthesized and transported in the phloem of higher plants. The polymer-trapping mechanism proposed for dicotyledonous C3 species may also be valid for oligofructan transport in monocotyledonous species, such as A. deserti, which may use a symplastic pathway for phloem loading of photosynthates in its mature leaves.

Journal Article↗

PubMed1998

Serial observations after high dose talc slurry in the rabbit model for pleurodesis.

The mechanisms leading to a pleurodesis after the intrapleural injection of a sclerosing agent are not completely understood. The purpose of the present study was to make serial observations over 28 days on the pleural fluid findings and the gross and microscopic changes in the pleura after talc slurry administered intrapleurally at a high dose. Sixty-six rabbits received 400 mg/kg talc slurry. Ten to 12 rabbits were sacrificed 1, 2, 4, 7, 14, and 28 days after the intrapleural injection. At sacrifice the pleural fluid was measured and analyzed, and the pleural surfaces were studied grossly and microscopically. The intrapleural injection of 400 mg/kg talc slurry resulted in an acute exudative pleural effusion that persisted for 4 days. There was a progressive increase in the gross and microscopic fibrosis over the 28 days. Talc was present at the time of sacrifice in all animals. At 28 days there was a clinically significant pleurodesis in all rabbits; pleurodesis was not observed before this time. From this study we conclude that the intrapleural injection of 400 mg/kg talc slurry leads to an acute exudative pleural effusion and clinically significant pleurodesis that is present on day 28 but not day 14. It appears that the production of a pleurodesis requires higher doses of talc in rabbits without a chest tube than in humans with a chest tube.

Acute Disease↗

PubMed1998

The effect of antibody against vascular endothelial growth factor on tumor growth and metastasis.

Vascular endothelial growth factor (VEGF), a very important in the process of tumor angiogenesis, was chosen as a target in a study to determine whether manipulation of angiogenesis with antibody against VEGF may interrupt tumor growth and metastasis. Anti-VEGF antibody was obtained from immunized rabbits, purified on an affinity column, and identified as neutralized antibody by Mile's assay. IVTA2MA891, a murine spontaneous breast cancer with a high rate of metastasis in lung in TA2 x 615 F1 mice, was chosen as an animal model in this study, because of the high expression of VEGF in the primary tumor as well as in the lung metastatic tumor. The anti-VEGF antibody could inhibit growth of S180 sarcoma in a dose-dependent manner, and the inhibition rate could reach 41.0% with a dose of 200 microg mouse(-1) day(-1). Anti-VEGF antibody could inhibit tumor growth by 76.2% in nude mice bearing human gastric cancer (MGC 803). When anti-VEGF antibody was combined with 131I-3H11, a murine monoclonal antibody conjugated with 131I, only one of five nude mice developed tumor and 84.0% more inhibition of tumor growth was obtained in comparison with treatment by 131I-3H11 alone. The growth of the primary tumor was inhibited by 44.0% and the number and size of the metastatic foci in the lungs were reduced by 73.0% and 83.7% respectively in the animal model, with a high rate of metastasis in lung. The anti-VEGF antibody may be potentially useful for clinical treatment of cancer and metastasis.

Animals↗

PubMed1998

RT-PCR for tyrosinase-mRNA-positive cells in peripheral blood: evaluation strategy and correlation with known prognostic markers in 123 melanoma patients.

Reverse transcriptase polymerase chain reaction for the detection of tyrosinase-mRNA-positive cells in peripheral blood of melanoma patients, as a possible marker of hematogenous dissemination, has demonstrated varying detection rates. This study examined the sensitivity and reproducibility of the technique using a protocol of multiple polymerase chain reaction to determine circulating melanocytic cells. For each of the 123 melanoma patients included in this study, four nested polymerase chain reactions were performed from two blood specimens requiring both polymerase chain reactions from at least one blood sample to be positive to consider a patient as positive. Thus, a definitive result was obtained in 98% of the cases, whereas only 1.6% lacked conclusive findings. Thus, we found a correlation between the tyrosinase detection rate and the clinical stage. Circulating tyrosinase-mRNA-positive cells were detected in 13% of patients with primary tumor, 17% with regional skin/lymph node metastasis, and 44% with distant metastasis. Positivity also correlated with known melanoma progression markers such as gender, tumor thickness, and histologic type. Positive results were obtained more frequently in (i) men compared with women, (ii) patients with thick primary melanomas (> 4 mm: 38%) compared with those with thinner tumors (1.1-4 mm, 22%; < or = 1 mm, 5%), and (iii) patients with nonclassifiable (38%), nodular (34%), and occult primary melanomas (30%) compared with those with acrolentiginous (17%), superficial spreading (9%), or lentigo maligna melanoma (0%). These findings suggest that detection of tyrosinase-mRNA-positive cells in peripheral blood is not an adequate marker for identifying melanoma patients with distant metastasis. Reverse transcriptase polymerase chain positivity in early melanoma stages, however, as corresponding to other prognostic parameters, may indicate increased risk for the development of hematogenous metastasis and may be of value as a progression marker.

Adult↗

PubMed1998

Expression of a CD3 epsilon transgene in CD3 epsilon(null) mice does not restore CD3 gamma and delta expression but efficiently rescues T cell development from a subpopulation of prothymocytes.

The TCR-associated CD3 complex consists of four subunits, i.e. CD3 gamma, delta, epsilon and zeta, which are expressed very early in T cell development prior to the expression of the TCR and the pre-TCR alpha chain. It is unclear whether the expression of each CD3 protein is independent of, or is influenced by, other CD3 subunits. To study whether CD3 epsilon regulates expression of CD3 gamma and delta genes, we generated a strain of CD3 epsilon-deficient mice termed CD3 epsilon(delta P/delta P) (epsilon(delta P)), in which the promoter of CD3E was disrupted, and subsequently reconstituted these mice with a CD3 epsilon transgene. In the epsilon(delta P) mice, T cell development is arrested at the double-negative stage and targeting the CD3 epsilon gene caused severe inhibition of CD3 gamma and delta gene expression. Introduction of the CD3 epsilon transgene did not restore CD3 gamma and delta expression. However, a very small fraction of prothymocytes that expressed CD3 gamma and delta was rescued upon reconstitution of the CD3 epsilon transgene. Remarkably, this rescue led to a very efficient differentiation and maturation of thymocytes, resulting in a significant T cell population in the periphery. These results demonstrate that CD3 epsilon does not regulate expression of CD3 gamma and delta genes, and underscore the capacity of each prothymocyte to give rise to a large number of mature peripheral T cells.

Animals↗

PubMed1998

Prenatal identification of de novo marker chromosomes using micro-FISH approach.

Chromosome microdissection combined with polymerase chain reaction (PCR) and reverse chromosome painting ('micro-FISH') is a powerful technique for the unequivocal identification of complex or subtle chromosomal aberrations. We have applied this technique to the prenatal diagnosis of three fetuses with de novo marker chromosomes. One small supernumerary satellited marker chromosome was shown to have originated from the fusion of the centromeric heterochromatin of one or both of chromosomes 14 and 22. The second marker was identified as i(9)(p10) while the third marker chromosome was shown to have been derived from the 1p13.1-1q21.3 region. At birth, the clinical outcome correlated well with that expected from the prenatal cytogenetic findings. Our study highlights the importance of the application of 'micro-FISH' to prenatal diagnosis.

Adolescent↗

PubMed1998

Is cytoskeletal tension a major determinant of cell deformability in adherent endothelial cells?

We tested the hypothesis that mechanical tension in the cytoskeleton (CSK) is a major determinant of cell deformability. To confirm that tension was present in adherent endothelial cells, we either cut or detached them from their basal surface by a microneedle. After cutting or detachment, the cells rapidly retracted. This retraction was prevented, however, if the CSK actin lattice was disrupted by cytochalasin D (Cyto D). These results confirmed that there was preexisting CSK tension in these cells and that the actin lattice was a primary stress-bearing component of the CSK. Second, to determine the extent to which that preexisting CSK tension could alter cell deformability, we developed a stretchable cell culture membrane system to impose a rapid mechanical distension (and presumably a rapid increase in CSK tension) on adherent endothelial cells. Altered cell deformability was quantitated as the shear stiffness measured by magnetic twisting cytometry. When membrane strain increased 2.5 or 5%, the cell stiffness increased 15 and 30%, respectively. Disruption of actin lattice with Cyto D abolished this stretch-induced increase in stiffness, demonstrating that the increased stiffness depended on the integrity of the actin CSK. Permeabilizing the cells with saponin and washing away ATP and Ca2+ did not inhibit the stretch-induced stiffening of the cell. These results suggest that the stretch-induced stiffening was primarily due to the direct mechanical changes in the forces distending the CSK but not to ATP- or Ca(2+)-dependent processes. Taken together, these results suggest preexisting CSK tension is a major determinant of cell deformability in adherent endothelial cells.

Cell Adhesion↗

PubMed1998

Regulation of cytoskeletal mechanics and cell growth by myosin light chain phosphorylation.

The role of myosin light chain phosphorylation in regulating the mechanical properties of the cytoskeleton was studied in NIH/3T3 fibroblasts expressing a truncated, constitutively active form of smooth muscle myosin light chain kinase (tMK). Cytoskeletal stiffness determined by quantifying the force required to indent the apical surface of adherent cells showed that stiffness was increased twofold in tMK cells compared with control cells expressing the empty plasmid (Neo cells). Cytoskeletal stiffness quantified using magnetic twisting cytometry showed an approximately 1.5-fold increase in stiffness in tMK cells compared with Neo cells. Electronic volume measurements on cells in suspension revealed that tMK cells had a smaller volume and are more resistant to osmotic swelling than Neo cells. tMK cells also have smaller nuclei, and activation of mitogen-activated protein kinase (MAP kinase) and translocation of MAP kinase to the nucleus are slower in tMK cells than in control cells. In tMK cells, there is also less bromodeoxyuridine incorporation, and the doubling time is increased. These data demonstrate that increased myosin light chain phosphorylation correlates with increased cytoskeletal stiffness and suggest that changing the mechanical characteristics of the cytoskeleton alters the intracellular signaling pathways that regulate cell growth and division.

3T3 Cells↗

PubMed1998

Extracellular matrix and pulmonary hypertension: control of vascular smooth muscle cell contractility.

Pulmonary hypertension is characterized by increased vascular resistance due to smooth muscle cell hyper-activity and excess deposition of extracellular matrix (ECM) in the vessel wall. We investigated the possibility that changes in cell-ECM interactions may play an active role in this process by modifying the contractile response of pulmonary vascular smooth muscle (PVSM) cells. Contractility was measured within individual cultured PVSM cells, when resting or stimulated with vasoactive agents, by quantitating changes in stiffness of the cytoskeleton (CSK) using magnetic twisting cytometry (N. Wang, J. P. Butler, and D. E. Ingber. Science 260: 1124-1127, 1993). Control studies confirmed that changes in CSK stiffness closely paralleled alterations in cell contraction and relaxation as measured in response to endothelin-1 (ET-1) and dibutyryl guanosine 3',5'-cyclic monophosphate (cGMP), respectively, in a collagen gel contraction assay. CSK stiffness and contractile tone in cultured PVSM cells increased in direct proportion as the density of fibronectin (FN) coating was raised from 10 to 500 ng/well in 96-well plates. Dibutyryl cGMP had no effect in cells on low FN, although it completely inhibited the FN-dependent increase in CSK stiffness on higher ECM densities. In contrast, ET-1 induced the greatest increase in CSK stiffness on the intermediate FN density (100 ng/well). The reduced sensitivity to ET-1 on high FN was not due to dysfunction of the contractile apparatus nor to changes in protein tyrosine phosphorylation. Taken together, these results show that ECM can modulate PVSM cell contractility and suggest that the changes in ECM observed in hypertensive vessels could play an important role in the etiology of this disease.

Animals↗

PubMed1998

Effect of renin gene transfer on blood pressure in the spontaneously hypertensive rat.

To investigate whether molecular variation in the renin gene contributes to the greater blood pressure of spontaneously hypertensive rats (SHR) versus normotensive Brown Norway (BN) rats, we measured blood pressure in an SHR progenitor strain and an SHR congenic strain that are genetically identical except at the renin gene and an associated segment of chromosome 13 transferred from the BN strain. Backcross breeding and molecular selection at the renin locus were used to create the SHR congenic strain (designated SHR.BN-Ren) that carries the renin gene transferred from the normotensive BN strain. We found that transfer of the renin gene from the BN strain onto the genetic background of the SHR did not decrease blood pressure in rats fed either a normal or high-salt diet. In fact, the systolic blood pressures of the SHR congenic rats tended to be slightly greater than the systolic blood pressures of the SHR progenitor rats. However, the congenic strain exhibited lower serum high-density lipoprotein cholesterol, and greater levels of total cholesterol, very-low-density lipoprotein, and intermediate-density lipoprotein cholesterol during administration of a high-fat, high-cholesterol diet. These findings demonstrate that (1) under the environmental circumstances of the current study, the greater blood pressure of SHR versus BN rats cannot be explained by strain differences in the renin gene and (2) a quantitative trait locus affecting lipid metabolism exists on chromosome 13 within the transferred chromosome segment. The SHR.BN-Ren congenic strain may provide a useful new animal model for studying the interaction between high blood pressure and dyslipidemia in cardiovascular disease.

Animals↗

PubMed1998

Mechanical interactions among cytoskeletal filaments.

Mechanical properties of the cells are important in controlling cell shape, cell migration, and other functions. To understand how cytoskeletal (CSK) filaments interact with one another mechanically, mechanical properties of adherent endothelial cells were analyzed after treatment with CSK-disrupting drugs. CSK stiffness (the ratio of applied stress to strain, a measure of cell resistance to shape deformation), viscosity (an index of intracellular structural damping), and permanent deformation (a measure of "plasticity") were measured with magnetic twisting cytometry, by which rotational stress was applied directly to integrin receptors with ferromagnetic beads coated with RGD-containing peptide. Treatment with cytochalasin D, which disrupts actin microfilaments inhibited stiffness by 50% and decreased permanent deformation from 70% to 50% but had almost no effect on viscosity. In contrast, nocodazole, a microtubule disrupter, had very little effect on inhibition of CSK stiffness, decreased viscosity by 25%, and had no effects on permanent deformation. Acrylamide, an intermediate filament disrupter, had little effect on inhibition of CSK stiffness, little effect on viscosity, and no effect on permanent deformation. Taxol, a drug that facilitates microtubule polymerization, increased stiffness by 10%, increased viscosity by 10%, and decreased permanent deformation from 70% to 50%. Combinations of cytochalasin D and nocodazole, cytochalasin D and acrylamide, or all three drugs resulted in a synergistic effect on inhibition of CSK stiffness and viscosity but not in permanent deformation. Inhibition of oxidative metabolism with potassium cyanide had no effects on stress-induced stiffening response. Inhibition of tyrosine phosphatase with phenylarsine oxide had no effect on stress-induced stiffening response. We conclude that higher order mechanical interactions of CSK filaments are important in determining the mechanical properties of the cell.

Acrylamide↗

PubMed1998

ACE, MTHFR, factor V Leiden, and APOE polymorphisms in patients with vascular and Alzheimer's dementia.

BACKGROUND AND PURPOSE: There is a growing interest in the use of genetic markers in the differential diagnosis of dementia. In the current study we examined the usefulness of genetic risk factors for vascular disease as markers for vascular dementia (VD). METHODS: The groups included 41 patients with VD, 49 patients with dementia of the Alzheimer's type, and 40 age-matched control subjects without dementia. These patients were genotyped for vascular disease-associated polymorphisms in the genes coding for methylenetetrahydrofolate reductase (MTHFR), angiotensin-converting enzyme (ACE), factor V Leiden (FVL), and a common genetic risk factor for AD, apolipoprotein E epsilon4 (APOE epsilon4). RESULTS: There was no significant association between ACE, MTHFR, and FVL genotypes with VD whether compared with subjects with AD or with control subjects. There was a higher frequency of APOE epsilon4 alleles in patients with AD (30%, P=0.016) and VD (26%, P=0.07) compared with control subjects (15%). CONCLUSIONS: VD is not associated with the genetic risk factors for vascular disease examined in this study, indicating that the pathogenesis of VD may differ from other vascular diseases.

Aged↗

PubMed1998

Impaired mechanical stability, migration and contractile capacity in vimentin-deficient fibroblasts.

Loss of a vimentin network due to gene disruption created viable mice that did not differ overtly from wild-type littermates. Here, primary fibroblasts derived from vimentin-deficient (-/-) and wild-type (+/+) mouse embryos were cultured, and biological functions were studied in in vitro systems resembling stress situations. Stiffness of -/- fibroblasts was reduced by 40% in comparison to wild-type cells. Vimentin-deficient cells also displayed reduced mechanical stability, motility and directional migration towards different chemo-attractive stimuli. Reorganization of collagen fibrils and contraction of collagen lattices were severely impaired. The spatial organization of focal contact proteins, as well as actin microfilament organization was disturbed. Thus, absence of a vimentin filament network does not impair basic cellular functions needed for growth in culture, but cells are mechanically less stable, and we propose that therefore they are impaired in all functions depending upon mechanical stability.

Actins↗

PubMed1998

[One stage repair and reconstruction for severe deep burns with compound tissue defects of upper limb].

In order to solve the difficult problems of repair and reconstruction for severe deep burns with compound tissue defects of upper limb, 26 cases were treated with transplantation of compound tissue flap, vascularized by anastomosis of blood vessel or by vascular pedicle. Several kinds of reparative and reconstructive procedure could be performed simultaneously. Not only the tissue defect was repaired, but also the upper limb function was reconstructed in one stage operation. Owing to the presence of abundant vascular supply from the vascularized compound tissue and primarily closing the wounds, the anti-infection potency was high, then it was suitable for such conditions as fresh severe deep burn with infection and compound tissue defects. As a result, this technique provided the best chance to save upper limb from amputation. The duration required for treatment could be markedly shortened. All the cases successed. The long-term functional recovery was satisfactory. This method provided the possibility to solve effectively the difficult problem dealing with the severe deep burns with compound tissue defects of upper limb.

Adolescent↗

PubMed1998

[Restoration of elbow joint and its function by transposition of latissimus dorsi myocutaneous flap following serious high-voltage injury in upper limb].

OBJECTIVE: This is to introduce a method for repairing and restoring the elbow joints and their function of the patients suffering from serious high-voltage electric burns in upper extremity. METHODS: We designed the transposition of latissimus dorsi myocutaneous flap (MCF) to repair the wounds of elbow, as well as restore the joint's flexion function by the motor power of the muscle. This procedure salvaged the elbow joints, which would have to be amputated before. RESULTS: 8 cases, 10 limbs were involved. All flaps survived completely and 6 of them had primary healing while the other 4 flaps healed after dressing changing. The elbow joints and their functions were satisfactorily restored in 5 limbs and the functions were also regained through further elbow joint release procedures in 2 limbs. The other 3 limbs (2 cases) lost contact with us and so the results were not clear. CONCLUSION: This method has evident advantages and satisfactory results.

Adult↗