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991.
Theurillat JP Zürrer-Härdi U Varga Z Storz M Probst-Hensch NM Seifert B Fehr MK Fink D Ferrone S Pestalozzi B Jungbluth AA Chen YT Jäger D Knuth A Moch H 《Cancer immunology, immunotherapy : CII》2007,56(11):1723-1731
NY-BR-1 is a recently identified differentiation antigen of the mammary gland. To use NY-BR-1 for T-cell-based immunotherapy, analysis of its co-expression with HLA class I antigens is required. In the present tissue microarray study, primary breast cancers (n = 1,444), recurrences (n = 88), lymph node (n = 525) and distant metastases (n = 91) were studied for NY-BR-1 expression using a novel monoclonal antibody. NY-BR-1 expression was compared with prognosis, estrogen receptor, HER2-status, EGFR and HLA class I antigen expression. NY-BR-1 was more frequently expressed in grade 1 (82%) than in grade 2 (69%) and grade 3 (46%) carcinomas (P < 0.0001). Moreover, NY-BR-1 expression correlated directly with estrogen receptor expression (P < 0.0001) and inversely correlated with HER2-status and EGFR expression (P < 0.0001 for both). Considering high expression level of co-expression, 198/1,321 (15%) primary breast carcinomas and 4/65 (6%) distant metastases expressed NY-BR-1 and HLA class I, suggesting that active immunotherapy can be applied to about 10% of breast cancer patients. Survival analysis showed an association of NY-BR-1 expression with better patient outcome (P = 0.015). No difference between NY-BR-1 expression of primary tumors and metastases could be found, indicating that the presence of NY-BR-1 in metastases can be deduced from their corresponding primary. Forty-three paired biopsies taken from patients before and after chemotherapy suggest that NY-BR-1 expression is not influenced by preceding chemotherapy (kappa = 0.89, P < 0.0001). In summary, the co-expression of NY-BR-1 with HLA class I antigens and its expression in metastases without modification by chemotherapy suggest that NY-BR-1 targeted immunotherapy represents a viable strategy in addition to other targeted cancer drug therapies of breast cancer. 相似文献
992.
Watts SW Rondelli C Thakali K Li X Uhal B Pervaiz MH Watson RE Fink GD 《American journal of physiology. Heart and circulatory physiology》2007,292(5):H2438-H2448
Arterial remodeling occurs in response to mechanical and neurohumoral stimuli. We hypothesized that veins, which are not exposed to higher pressures in hypertension, would demonstrate less active remodeling than arteries. We assessed remodeling with two standard measures of arterial remodeling: vessel morphometry and the expression/function of matrix metalloproteinases (MMPs). Thoracic aorta and vena cava from sham normotensive and DOCA-salt hypertensive rats (110 +/- 4 and 188 +/- 8 mmHg systolic blood pressure, respectively) were used. Wall thickness was increased in DOCA-salt vs. sham aorta (301 +/- 23 vs. 218 +/- 14 mum, P < 0.05), as was medial area, but neither measure was altered in the vena cava. The aorta and vena cava expressed the gelatinases MMP-2, MMP-9, transmembrane proteinase MT1-MMP, and tissue inhibitor of metalloproteinase-2 (TIMP-2). Immunohistochemically, MMP-2 localized to smooth muscle in the aorta and densely in endothelium/smooth muscle of the vena cava. Western and zymographic analyses verified that MMP-2 was active in all vessels and less active in the vena cava than aorta. In hypertension, MMP-2 expression and activity in the aorta were increased (59.1 +/- 3.7 and 74.5 +/- 6.1 units in sham and DOCA, respectively, P < 0.05); similar elevations were not observed in the vena cava. MMP-9 was weakly expressed in all vessels. MT1-MMP was expressed by the aorta and vena cava and elevated in the vena cava from DOCA-salt rats. TIMP-2 expression was significantly increased in the aorta of DOCA rats compared with sham but was barely detectable in the vena cava of sham or DOCA-salt hypertensive rats. These findings suggest that large veins may not undergo vascular remodeling in DOCA-salt hypertension. 相似文献
993.
Microsensor studies on Padina from a natural CO2 seep: implications of morphology on acclimation to low pH
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Low seawater pH can be harmful to many calcifying marine organisms, but the calcifying macroalgae Padina spp. flourish at natural submarine carbon dioxide seeps where seawater pH is low. We show that the microenvironment created by the rolled thallus margin of Padina australis facilitates supersaturation of CaCO3 and calcifi‐cation via photosynthesis‐induced elevated pH. Using microsensors to investigate oxygen and pH dynamics in the microenvironment of P. australis at a shallow CO2 seep, we found that, under saturating light, the pH inside the microenvironment (pHME) was higher than the external seawater (pHSW) at all pHSW levels investigated, and the difference (i.e., pHME ? pHSW) increased with decreasing pHSW (0.9 units at pHSW 7.0). Gross photosynthesis (Pg) inside the microenvironment increased with decreasing pHSW, but algae from the control site reached a threshold at pH 6.5. Seep algae showed no pH threshold with respect to Pg within the pHSW range investigated. The external carbonic anhydrase (CA) inhibitor, acetazolamide, strongly inhibited Pg of P. australis at pHSW 8.2, but the effect was diminished under low pHSW (6.4–7.5), suggesting a greater dependence on membrane‐bound CA for the dehydration of HCO3? ions during dissolved inorganic carbon uptake at the higher pHSW. In comparison, a calcifying green alga, Halimeda cuneata f. digitata, was not inhibited by AZ, suggesting efficient bicarbonate transport. The ability of P. australis to elevate pHME at the site of calcification and its strong dependence on CA may explain why it can thrive at low pHSW. 相似文献
994.
Lillie Cavonius Helen Fink Juris Kiskis Eva Albers Ingrid Undeland Annika Enejder 《Plant physiology》2015,167(3):603-616
Microalgae have great prospects as a sustainable resource of lipids for refinement into nutraceuticals and biodiesel, which increases the need for detailed insights into their intracellular lipid synthesis/storage mechanisms. As an alternative strategy to solvent- and label-based lipid quantification techniques, we introduce time-gated coherent anti-Stokes Raman scattering (CARS) microscopy for monitoring lipid contents in living algae, despite strong autofluorescence from the chloroplasts, at approximately picogram and subcellular levels by probing inherent molecular vibrations. Intracellular lipid droplet synthesis was followed in Phaeodactylum tricornutum algae grown under (1) light/nutrient-replete (control [Ctrl]), (2) light-limited (LL), and (3) nitrogen-starved (NS) conditions. Good correlation (r2 = 0.924) was found between lipid volume data yielded by CARS microscopy and total fatty acid content obtained from gas chromatography-mass spectrometry analysis. In Ctrl and LL cells, micron-sized lipid droplets were found to increase in number throughout the growth phases, particularly in the stationary phase. During more excessive lipid accumulation, as observed in NS cells, promising commercial harvest as biofuels and nutritional lipids, several micron-sized droplets were present already initially during cultivation, which then fused into a single giant droplet toward stationary phase alongside with new droplets emerging. CARS microspectroscopy further indicated lower lipid fluidity in NS cells than in Ctrl and LL cells, potentially due to higher fatty acid saturation. This agreed with the fatty acid profiles gathered by gas chromatography-mass spectrometry. CARS microscopy could thus provide quantitative and semiqualitative data at the single-cell level along with important insights into lipid-accumulating mechanisms, here revealing two different modes for normal and excessive lipid accumulation.The accumulation of lipids in microalgae is currently a field of intense research: with their high photosynthetic efficiency and rapid growth rates, these organisms hold great potential both for sustainable production of biofuels (Chisti, 2007) and as a nutrition source (de Jesus Raposo et al., 2013). As in all living cells, lipids in microalgae are present in membranes, such as the plasma and organelle membranes. Some microalgae also accumulate lipids, mainly triacylglycerols, in intracellular droplets (De Martino et al., 2011; White et al., 2012). One such microalga is Phaeodactylum tricornutum, a unicellular photoautotrophic diatom and a well-studied model organism. It has a sequenced genome and is known to produce long-chain n-3 polyunsaturated fatty acids (PUFAs), eicosapentaenoic acid (EPA), and small amounts of docosahexaenoic acid (DHA; Alonso et al., 1998). The long-chain n-3 PUFA-producing properties has made it particularly interesting within the areas of functional food and nutraceuticals. Under conditions of nitrogen starvation, P. tricornutum accumulates larger amounts of fatty acids, albeit of the more saturated nature (Yongmanitchai and Ward, 1991). However, in response to low irradiance, P. tricornutum has been reported to increase its content of PUFAs, especially EPA (Thompson et al., 1990). In order to optimize strain selection and algal cultivation conditions in relation to lipid accumulation/lipid profile, accurate tools to quantify total lipids as well as the degree of unsaturation are required.Solvent extractions followed by methylation, gas chromatography coupled to flame ionization detection, and gas chromatography-mass spectrometry (GC-MS) detection belong to the standard techniques used for algal lipid analysis. However, these are cumbersome and require relatively large amounts of solvents and sample. The resulting quantitative information on lipid amounts is related to total cell mass, which may introduce artifacts, as the cell mass is also influenced by other metabolic parameters. Furthermore, these bioanalytical techniques provide rough population averages without information on the intracellular location or distribution. As an alternative, microscopy techniques are increasingly being used, with the benefit that lipid droplets can be evaluated directly in single cells with high precision. Fluorescence microscopy is the most widespread technique, relying on lipid-specific fluorescent markers: Nile Red is commonly used, but its poor permeability through the cell walls causes difficulties, as does its nonspecific binding (Chen et al., 2009). Other fluorophores like BODIPY 505/515 also have been suggested for live-cell studies (Cooper et al., 2010; Wong and Franz, 2013). Still, invasive techniques are needed, requiring solvents to facilitate the transport of the labeling molecules into the cell, potentially inducing stress responses that may affect cell metabolism. Furthermore, there is little knowledge available on how the accumulation of lipophilic dyes in lipid droplets and the fluorescence emission are influenced by environmental conditions such as temperature, pH, and deposited light doses. It has also been shown that the fluorescence intensity emitted from the dyes cannot be related directly to the local lipid concentration, excluding quantitative measurements (De la Hoz Siegler et al., 2012). In algae/plant cell biology, the applicability of fluorescence microscopy is also limited due to the fact that algae/plant cells generate strong autofluorescence, potentially interfering with the lipid/fatty acid signals. In order to take algal lipid quantification one step further, microscopy techniques, not being dependent on exogenous fluorophores and allowing efficient separation of the lipid/fatty acid signal from the autofluorescence, are desirable.In label-free coherent anti-Stokes Raman scattering (CARS) microscopy, images are formed by probing intrinsic molecular vibrations through a nonlinear four-wave-mixing process (Cheng and Xie, 2004). Briefly, the frequency difference of two coherent near-infrared excitation beams (the pump beam at shorter wavelengths and the Stokes beam at longer wavelengths) are tuned to match the frequency of the target molecular vibration. As a result, resonant oscillators are coherently driven in the sample focal volume and an enhanced blue-shifted CARS signal is generated. Due to the nonlinear nature of the CARS process, emission is generated only in the high-intensity region of focused laser beams, allowing optical sectioning of the specimen. By tuning the frequency difference of the excitation beams to match the resonance frequency of carbon-hydrogen (C-H) vibrations, especially abundant in lipids, three-dimensional images of lipids can be recorded without any staining (Enejder et al., 2010). As the CARS emission scales with the square of the concentration of C-H bonds, quantitative data on intracellular amounts of lipids can be extracted (Cheng and Xie, 2004). However, cells with chromophores, such as algae and plant cells, generate exceptionally strong two-photon fluorescence, the spectral tails of which tend to bleed through the most efficient optical filters typically used for separation of the CARS signal. As an alternative approach, we have incorporated a time-correlated single-photon counting system (Enejder et al., 2010), allowing us to distinguish the long-decay fluorescence component from the instant CARS signal by time gating.The capability of conventional CARS imaging for microalgae was recently demonstrated with proof-of-principle data, showing that individual, larger lipid droplets can be resolved visually, in contrast to conventional Raman microscopy (He et al., 2012). In this study, we introduce CARS microscopy with time-gated detection, also enabling the identification of subpicogram lipid-rich regions in the vicinity of strongly autofluorescent chloroplasts. This is particularly important because cellular storage lipids in algae are primarily synthesized in the chloroplasts and then budded off from the envelope membranes as nascent lipid droplets (Fan et al., 2011). Hence, time-gated CARS microscopy paves the way for high-precision quantification of the complete intracellular distribution of lipid stores, including the emerging droplets within and adjacent to chloroplasts. We show the feasibility of the approach for quantitative lipid analysis of large populations of living P. tricornutum cultivated under three different growth conditions: a control (Ctrl) group, a light-limited (LL) group to increase the EPA level, and a nitrogen-starved (NS) group to increase total lipid accumulation. We studied the lipid metabolism in approximately 100 cells per category over time and report detailed visual information on the dynamics of lipid droplet formation throughout a life cycle from budding droplets to, in some cases, giant lipid stores. We show that biologically relevant quantitative and qualitative data on intracellular lipid stores can be extracted at a precision of less than 1 pg cell−1 despite adjacent chromophores. Data extracted from the CARS images are further compared with solvent extraction and quantification of fatty acids using GC-MS. We further illustrate the potential to assess whether the different growth conditions promote the synthesis of more PUFAs by detecting shifts in lipid fluidity and saturation per individual lipid droplet from CARS
C-H vibration ratio images. 相似文献
995.
Ssy1p and Ptr3p Are Plasma Membrane Components of a Yeast System That Senses Extracellular Amino Acids
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Mutations in SSY1 and PTR3 were identified in a genetic selection for components required for the proper uptake and compartmentalization of histidine in Saccharomyces cerevisiae. Ssy1p is a unique member of the amino acid permease gene family, and Ptr3p is predicted to be a hydrophilic protein that lacks known functional homologs. Both Ssy1p and Ptr3p have previously been implicated in relaying signals regarding the presence of extracellular amino acids. We have found that ssy1 and ptr3 mutants belong to the same epistasis group; single and ssy1 ptr3 double-mutant strains exhibit indistinguishable phenotypes. Mutations in these genes cause the nitrogen-regulated general amino acid permease gene (GAP1) to be abnormally expressed and block the nonspecific induction of arginase (CAR1) and the peptide transporter (PTR2). ssy1 and ptr3 mutations manifest identical differential effects on the functional expression of multiple specific amino acid transporters. ssy1 and ptr3 mutants have increased vacuolar pools of histidine and arginine and exhibit altered cell growth morphologies accompanied by exaggerated invasive growth. Subcellular fractionation experiments reveal that both Ssy1p and Ptr3p are localized to the plasma membrane (PM). Ssy1p requires the endoplasmic reticulum protein Shr3p, the amino acid permease-specific packaging chaperonin, to reach the PM, whereas Ptr3p does not. These findings suggest that Ssy1p and Ptr3p function in the PM as components of a sensor of extracellular amino acids. 相似文献
996.
Ajith Rajapaksha Michael Fink Brian A. Todd 《Molecular & cellular biomechanics : MCB》2015,12(3):215-230
Delivery of therapeutic agents to the eye requires efficient transport through cellular and extracellular barriers. We evaluated the rate of diffusive transport in excised porcine corneal stroma using fluorescently labeled dextran molecules with hydrodynamic radii ranging from 1.3 to 34 nm. Fluorescence correlation spectroscopy (FCS) was used to measure diffusion coefficients of dextran molecules in the excised porcine corneal stroma. The preferential sensitivity of FCS to diffusion along two dimensions was used to differentially probe diffusion along the directions parallel to and perpendicular to the collagen lamellae of the corneal stroma. In order to develop an understanding of how size affects diffusion in cornea, diffusion coefficients in cornea were compared to diffusion coefficients measured in a simple buffer solution. Dextran molecules diffuse more slowly in cornea as compared to buffer solution. The reduction in diffusion coefficient is modest however (67% smaller), and is uniform over the range of sizes that we measured. This indicates that, for dextrans in the 1.3 to 34 nm range, the diffusion landscape of corneal stroma can be represented as a simple liquid with a viscosity approximately 1.5 times that of water. Diffusion coefficients measured parallel vs. perpendicular to the collagen lamellae were indistinguishable. This indicates that diffusion in the corneal stroma is not highly anisotropic. Our results support the notion that the corneal stroma is highly permeable and isotropic to transport of hydrophilic molecules and particles with hydrodynamic radii up to at least 34 nm. 相似文献
997.
Chronic stretch of engineered heart tissue induces hypertrophy and functional improvement. 总被引:13,自引:0,他引:13
To examine the influence of chronic mechanical stretch on functional behavior of cardiac myocytes, we reconstituted embryonic chick or neonatal rat cardiac myocytes to a 3-dimensional engineered heart tissue (EHT) by mixing freshly isolated cells with neutralized collagen I and culturing them between two Velcro-coated silicone tubes, held at a fixed distance with a metal spacer. After 4 days, EHTs were subjected to a phasic unidirectional stretch for 6 days in serum-containing medium. Compared to unstretched controls, RNA/DNA and protein/cell ratios increased by 100% and 50%, respectively. ANF mRNA and alpha-sarcomeric actin increased by 98% and 40%, respectively. Morphologically, stretched EHTs exhibited improved organization of cardiac myocytes into parallel arrays of rod-shaped cells, increased cell length and width, longer myofilaments, and increased mitochondrial density. Thus, stretch induced phenotypic changes, generally referred to as hypertrophy. Concomitantly, force of contraction was two- to fourfold higher both under basal conditions and after stimulation with calcium or the beta-adrenergic agonist isoprenaline. Contraction kinetics were accelerated with a 14-44% decrease in twitch duration under all those conditions. In summary, we have developed a new in vitro model that allows morphological, molecular, and functional consequences of stretch to be studied under defined conditions. The main finding was that stretch of EHTs induced cardiac myocyte hypertrophy, which was accompanied by marked improvement of contractile function. 相似文献
998.
Sierra N. Clark James E. Bennett Raphael E. Arku Allan G. Hill Günther Fink Richard M. Adanu Richard B. Biritwum Rudolph Darko Ayaga Bawah Rosemary B. Duda Majid Ezzati 《PLoS medicine》2021,18(11)
BackgroundBody-mass index (BMI) and blood pressure (BP) levels are rising in sub-Saharan African cities, particularly among women. However, there is very limited information on how much they vary within cities, which could inform targeted and equitable health policies. Our study aimed to analyse spatial variations in BMI and BP for adult women at the small area level in the city of Accra, Ghana.Methods and findingsWe combined a representative survey of adult women’s health in Accra, Ghana (2008 to 2009) with a 10% random sample of the national census (2010). We applied a hierarchical model with a spatial term to estimate the associations of BMI and systolic blood pressure (SBP) and diastolic blood pressure (DBP) with demographic, socioeconomic, behavioural, and environmental factors. We then used the model to estimate BMI and BP for all women in the census in Accra and calculated mean BMI, SBP, and DBP for each enumeration area (EA). BMI and/or BP were positively associated with age, ethnicity (Ga), being currently married, and religion (Muslim) as their 95% credible intervals (95% CrIs) did not include zero, while BP was also negatively associated with literacy and physical activity. BMI and BP had opposite associations with socioeconomic status (SES) and alcohol consumption. In 2010, 26% of women aged 18 and older had obesity (BMI ≥ 30 kg/m2), and 21% had uncontrolled hypertension (SBP ≥ 140 and/or DBP ≥ 90 mm Hg). The differences in mean BMI and BP between EAs at the 10th and 90th percentiles were 2.7 kg/m2 (BMI) and in BP 7.9 mm Hg (SBP) and 4.8 mm Hg (DBP). BMI was generally higher in the more affluent eastern parts of Accra, and BP was higher in the western part of the city. A limitation of our study was that the 2010 census dataset used for predicting small area variations is potentially outdated; the results should be updated when the next census data are available, to the contemporary population, and changes over time should be evaluated.ConclusionsWe observed that variation of BMI and BP across neighbourhoods within Accra was almost as large as variation across countries among women globally. Localised measures are needed to address this unequal public health challenge in Accra.Sierra N. Clark and colleagues analyze spatial variations of blood pressure and body mass index, and associated factors in Ghanaian women. 相似文献
999.
Physiological responses to stoichiometric constraints: nutrient limitation and compensatory feeding in a freshwater snail 总被引:6,自引:0,他引:6
Nitrogen (N) and phosphorus (P) are considered to be essential nutrients that control secondary production in various ecosystems; insufficient availability of N and P can limit herbivore growth. Here, data are presented from field samplings and from a laboratory experiment on the potential of primary producers low in P, N, or P and N to constrain growth of the freshwater gastropod Radix ovata . The filamentous green alga Ulothrix fimbriata was cultured under different nutrient regimes, resulting in algae with different C:N:P ratios. The pure algae were fed in high and low quantities to juvenile R. ovata . Low availability of N and especially P in the algae strongly constrained the biomass accrual of the herbivore. In accordance with theoretical predictions, these food quality differences were highly dependent on the food quantity. The snails' growth rate was significantly related to their body C:P ratio, thereby supporting the growth rate hypothesis. R. ovata displayed a pronounced compensatory feeding response to low-nutrient food that could partly dampen but not fully compensate the food quality effects on snail growth. Increased feeding of gastropods at low P and/or N availability leads to depletion of periphyton biomass; hence compensatory feeding would shift the benthic herbivore community from a P or N limitation to a C limitation and thus have whole-ecosystem effects. 相似文献
1000.
Levy RM Mollen KP Prince JM Kaczorowski DJ Vallabhaneni R Liu S Tracey KJ Lotze MT Hackam DJ Fink MP Vodovotz Y Billiar TR 《American journal of physiology. Regulatory, integrative and comparative physiology》2007,293(4):R1538-R1544
High-mobility group box 1 (HMGB1) is a 30-kDa DNA-binding protein that displays proinflammatory cytokine-like properties. HMGB1-dependent inflammatory processes have been demonstrated in models of sterile injury, including ischemia-reperfusion injury and hemorrhagic shock. Here, we tested the hypothesis that the systemic inflammatory response and associated remote organ injury that occur after peripheral tissue injury are highly dependent on HMGB1. Toll-like receptor 4 (TLR4) wild-type (WT) mice subjected to bilateral femur fracture after treatment with neutralizing antibodies to HMGB1 had lower serum IL-6 and IL-10 levels compared with mice treated with nonimmune control IgG. Similarly, compared with injured mice treated with control IgG, anti-HMGB1 antibody-treated mice had lower serum alanine aminotransferase levels and decreased hepatic and gut mucosal NF-kappaB DNA binding. TLR4 mutant (C3H/HeJ) mice subjected to bilateral femur fracture had less systemic inflammation and liver injury than WT controls. Residual trauma-induced systemic inflammation and hepatocellular injury were not ameliorated by treatment with a polyclonal anti-HMGB1 antibody, even though HMGB1 levels were transiently elevated just 1 h after injury in both WT and C3H/HeJ mice. Collectively, these data demonstrate a critical role for a TLR4-HMGB1 pathway in the initiation of systemic inflammation and end-organ injury following isolated peripheral tissue injury. 相似文献