Modeling of intraorgan arterial vasculature. II. Propagation of pressure waves

The dependence of the wave conductance in self-similar dichotomous models of intraorgan arterial vasculatures on the model parameters was studied. It was found that, with different sets of parameters, it is possible to simulate the suction effect induced by negative reflections of waves from arterial branchings and to model the resonance properties of arterial beds. It was shown that the choice of an adequate model for a given intraorgan arterial vasculature should be based on agreement between the biophysical characteristics of the model and the bed that characterize the propagation and reflection of pulse waves.     

Modeling of intraorgan arterial vasculatures. I. A stationary blood flow at low Reynolds numbers

Based on the statistical relationships between the lengths and diameters of vessels in the arterial beds, obtained by measurements on plastic casts, a method for the generation of models of intraorgan arterial vasculatures was proposed. The dependence of the total hydraulic conductivity of the system on the model parameters was computed. The problem of selecting the adequate models by a comparison of the corresponding biophysical characteristics of vasculatures is discussed.     

Morphological adaptation of shape to flow: Microcurrents around lotic macroinvertebrates with known Reynolds numbers at quasi-natural flow conditions

Summary Using Laser Doppler Anemometry we measured current velocities in the median plane around dead lotic macroinvertebrates in a flume which reproduced natural near bottom hydraulics. We investigated specimens of the gastropods Ancylus, Acroloxus, and Potamopyrgus, the amphipod Gammarus, and the larval caddisflies Anabolia, Micrasema, and Silo of various size, various alignment to the flow or which were otherwise manipulated in order to clarify certain questions of adaptation of shape or case building style to flow, or the effects of flow on field distribution patterns. The steepest velocity gradients close to the animals were found near areas of their bodies protruding furthest into the flow. In such regions the rates of potential diffusive exchange processes, the potential corrasion (abrasion through suspended solids), and, for larger specimens, the lift forces (directed towards the water surface) must be highest. Posterior of these areas growing boundary layers formed above those species whose upper contour was approximately parallel to the upstream-downstream direction of the flow. All specimens removed momentum from the flow and thus experience a drag force (directed downstream). From the complete data set we derived the following general conclusions about the physical effects of potential morphological adaptations, taking into consideration diffusion through boundary layers, corrasion, lift forces, friction and pressure drag forces: The physical significance of these five factors generally depends on the Reynolds number of an animal and is largely affected by flow separation, which was significantly related to the ratio of body length to height and the slope of the posterior contour. A simultaneous effective morphological adaptation to all five factors is physically impossible and, in addition, would have to change from life at low (e.g. a young, small specimen of a species) to life at high (e.g. a fully grown specimen of the same species) Reynolds number.     

A low resting heart rate at diagnosis predicts favourable long-term outcome in pulmonary arterial and chronic thromboembolic pulmonary hypertension. A prospective observational study

Background

A low resting heart rate (HR) is prognostically favourable in healthy individuals and in patients with left heart disease. In this study we investigated the impact of HR at diagnosis on long-term outcome in patients with differently classified precapillary pulmonary hypertension (pPH).

Methods

pPH patients diagnosed as pulmonary arterial (PAH) or inoperable chronic thromboembolic pulmonary hypertension (CTEPH) were registered and regularly followed at our centre Baseline characteristics and events defined as either death or lung transplantation were noted. The prognostic value of HR was analysed using Kaplan Meier estimates, live tables and Cox regression.

Results

206 patients with PAH (148) and inoperable CTEPH (58) were included. The median HR was 82 bpm. pPH with a HR below 82 bpm had a significantly longer overall event-free survival (2409 vs.1332 days, p = .000). This advantage was similarly found if PAH and CTEPH were analysed separately. Although a lower HR was associated with a better hemodynamic and functional class, HR was a strong and independent prognostic marker for transplant free survival even if corrected for age, sex, hemodynamics and functional status.

Conclusion

We show that resting HR at diagnosis is a strong and independent long-term prognostic marker in PAH and CTEPH. Whether reducing HR by pharmacological agents would improve outcome in pPH has to be assessed by future trials with high attention to safety.     

Primary reactions of Photosystem II at low pH. I. Prompt and delayed fluorescence

Prompt and delayed chlorophyll fluorescence have been studied in broken spinach chloroplasts at pH values down to 2.6. No direct effect of low pH on the primary charge separation in Photosystem II was observed. The irreversible inactivation of a secondary electron donor in a narrow pH range around pH 4.5 was demonstrated. At lower pH values the photooxidized form of a more primary electron donor, revealed by its efficient fluorescence quenching, was reduced with a half time of about 200 μs, 25% by another electron donor and 75% by back reaction with the reduced acceptor. The electron donation had a half time of 800 μs and was practically irreversible. The back reaction had a pH dependent half time: about 270 μs at pH 4 and increasing towards lower pH. The competition of both reactions resulted in a net efficiency of the charge separation at pH 4 of 25%, increasing towards lower pH.     

Survival of Entamoeba and related Amoebae at low temperature. I. Viability of Entamoeba cysts at 4 degrees C

The time of survival at 4°C of mature cysts of six species of Entamoeba was studied. The length of time cysts generally survived storage was E. histolytica 12 weeks, E. moshkovskii 36 weeks, E. terrapinae and E. knowlesi 24 weeks, E. ranarum 25 weeks and E. invadens 7 weeks.     

Heparin-like glycosaminoglycans influence growth and phenotype of human arterial smooth muscle cells in vitro. I. Evidence for reversible binding and inactivation of the platelet-derived growth factor by heparin

Summary We have investigated the effects of interactions between growth factors and heparin-like glycosaminoglycans on untransformed human arterial smooth muscle cells (hASMC) in vitro. The results indicate that heparin in the presence of serum mitogens prevents the cells from entering the S phase of the cell cycle by binding and inactivating reversibly some serum mitogen(s). Our results suggest that platelet-derived growth factor (PDGF) is one of them and that it is the most potent stimulator of hASMC growth in vitro. Thymidine incorporation as well as increase in DNA content was inhibited not only by the presence of heparin in serum-containing medium but also when serum was chromatographed on Heparin-Sepharose at physiologic salt concentrations before exposure to the cells. The mitogenic activity of the unretained serum fraction was restored by the addition of PDGF AA, AB, or BB dimers or of a fraction (RF I) that dissociated from Heparin-Sepharose at 0.2 to 0.6M NaCl. Radiolabeled recombinant PDGF (c-sis) dissociated from Heparin-Sepharose within a concentration range of NaCl similar to that of RF I. Neither the unretained material nor the RF I or PDGF dimers were effective alone. The effect of RF I was significantly decreased by the addition of an anti-PDGF IgG that is known to neutralize the PDGF mitogenic activity partially. Addition of heparin abolished DNA-synthesis when the PDGF dimers or RF I were combined with the unretained fraction. A second fraction (RF II) bound strongly to Heparin-Sepharose and eluted between 1.1 and 1.6M NaCl. The RF II also induced DNA synthesis but was neither as efficient as RF I nor depending on other serum fractions for growth promotion and it was not inhibited by anti-PDGF IgG. A similar strong affinity for Heparin-Sepharose was found for labeled basic fibroblast growth factor and we cannot exclude the possibility that RF II represent fibroblast growth factor. Under these culture conditions, inhibition of hASMC proliferation was directly correlated with the expression of smooth muscle specific alpha actin isoforms in stress fibers and the suppression of a proliferating cell-specific nuclear antigen. Conversely, stimulation of hASMC proliferation was associated with the opposite phenomenon. We conclude that heparin-like glycosaminoglycans influence growth and phenotype of hASMCs in vitro by binding and inactivating PDGF. Inasmuch as heparin-like substances constitute a significant proportion of the proteoglycan-associated glycosaminoglycans of the arterial wall, such mechanisms might be important for the development of atherosclerotic lesions.     

Electrochromic absorbance changes of photosynthetic pigments in Rhodopseudomonas sphaeroides. I. Stimulation by secondary electron transport at low temperature

Light-induced absorbance changes were measured at temperatures between ?30 and ?55 °C in chromatophores of Rhodopseudomonas sphaeroides. Absorbance changes due to photooxidation of reaction center bacteriochlorophyll (P-870) were accompanied by a red shift of the absorption bands of a carotenoid. The red shift was inhibited by gramicidin D. The kinetics of P-870 indicated electron transport from the “primary” to a secondary electron acceptor. This electron transport was slowed down by lowering the temperature or increasing the pH of the suspension. Electron transport from soluble cytochrome c to P-870⁺ occurred in less purified chromatophore preparations. This electron transport was accompanied by a relatively large increase of the carotenoid absorbance change. This agrees with the hypothesis that P-870 is located inside the membrane, so that an additional membrane potential is generated upon transfer of an electron from cytochrome to P-870⁺.A strong stimulation of the carotenoid changes (more than 10-fold in some experiments) and pronounced band shifts of bacteriochlorophyll B-850 were observed upon illumination in the presence of artificial donor-acceptor systems. Reduced N-methylphenazonium methosulphate (PMS) and N,N,N′,N′-tetramethyl-p-phenylenediamine (TMPD) were fairly efficient donors, whereas endogenous ubiquinone and oxidized PMS acted as secondary acceptor. These results indicate the generation of large membrane potentials at low temperature, caused by sustained electron transport across the chromatophore membrane. The artificial probe, merocyanine MC-V did not show electrochromic band shifts at low temperature.     

Leaf conductances and xylem pressures of the host/mistletoe pair Eucalyptus behriana F. Muell. and Amyema miquelii (Lehm. ex Miq.) Tiegh. at permanently low plant water status in the field

Summary Over several days at permanently low plant water status in the field, where predawn xylem pressures () were never higher (less negative) than –1.2 MPa even after extended rain, leaf conductances (g) and transpiration rates of host trees, Eucalyptus behriana F. Muell., were higher than in mistletoes, Amyema miquelii (Lehm. ex Miq.) Tiegh., which contrasts with most studies known from the literature. Mistletoes influenced but not g of host leaves distal to the haustorium. Releasing xylem tension by cutting a host stem under water raised from about –3.5 MPa to about –0.5 MPa in both plants indicating that factors in the root zone were responsible for the low in the host. In all cases, with a freely transpiring or non-transpiring parasite at low and at artificially raised , mistletoe xylem pressure was lower than that of the host. Possible reasons are discussed.     

The site of photoinhibition in leaves of Cucumis sativus L. at low temperatures is photosystem I,not photosystem II

Maximum quantum yields (QY) of photosynthetic electron flows through PSI and PSII were separately assessed in thylakoid membranes isolated from leaves of Cucumis sativus L. (cucumber) that had been chilled in various ways. The QY(PSI) in the thylakoids prepared from the leaves treated at 4° C in moderate light at 220 mol quanta·m^–2·s^–1 (400–700 nm) for 5 h, was about 20–30% of that in the thylakoids prepared from untreated leaves, while QY(PSII) decreased, at most, by 20% in response to the same treatment. The decrease in QY(PSI) was observed only when the leaves were chilled at temperatures below 10° C, while such a marked temperature dependency was not observed for the decrease in QY(PSII). In the chilling treatment at 4° C for 5 h, the quantum flux density that was required to induce 50% loss of QY (PSI) was ca. 50 umol quanta·m^–2·s^–1. When the chilling treatment at 4° C in the light was conducted in an atmosphere of N₂, photoinhibition of PSI was largely suppressed, while the damage to PSII was somewhat enhanced. The ferricyanide-oxidised minus ascorbate-reduced difference spectra and the light-induced absorbance changes at 700 nm obtained with the thylakoid suspension, indicated the loss of P700 to extents that corresponded to the decreases in QY(PSI). Accordingly, the decreases in QY(PSI) can largely be attributed to destruction of the PSI reaction centre itself. These results clearly show that, at least in cucumber, a typical chillingsensitive plant, PSI is much more susceptible to aerobic photoinhibition than PSII.Abbreviations DCMU 3-(3,4-dichlorophenyl)-1,1-dimethylurea - P700 primary electron donor of PSI - PPFD photosynthetically active photon flux density - QY quantum yield We are grateful to invaluable comments by Prof. S. Katoh, K. Hikosaka and the members of our laboratory. We also thank A. Aoyama for technical assistance. This work was partly supported by the grants from the Ministry of Education, Science, and Culture, Japan, to I. Terashima (#03740342 and #04640621).     

Photofrin II Sensitized Modifications of Ion Transport Across the Plasma Membrane of an Epithelial Cell Line: I. Electrical Measurements at the Whole-Cell Level

Photofrin II is a photosensitizer frequently applied in photodynamic therapy. Light-induced tumor cell inactivation observed in the presence of this substance has been suggested to start with modifications at the level of cellular membranes. In the present study electrophysiological techniques are applied in order to investigate the action of photofrin II on functional properties of the plasma membrane of opossum kidney (OK) cells (as an epithelial model system) and of fibroblasts. Illumination of the cells in the presence of photofrin II (or Zn-phthalocyanine) leads to comparatively fast depolarization of the membrane potential. It is caused by a strong change of the membrane conductance which proceeds in two phases. Both phases contribute to a loss of ion selectivity of the plasma membrane between K⁺ and Na⁺. In the first phase, specific pathways for K⁺, which determine the resting potential under physiological conditions, are inactivated. The second phase is distinguished by a marked increase of a nonselective conductance. The increase of the latter — after light-induced initiation — continues in the dark. The conclusions are derived from light-induced, time-dependent changes of the membrane conductance and of the shape of the current-voltage relationship detected under different experimental conditions. Received: 26 May 1998/Revised: 8 September 1998     

Bovine oocytes treated prior to in vitro maturation with a combination of butyrolactone I and roscovitine at low doses maintain a normal developmental capacity.

Butyrolactone I (BL-I) and Roscovitine (ROS), two specific and potent inhibitors of M-phase promoting factor (MPF) kinase activity, were used to block germinal vesicle breakdown (GVBD) of cattle oocytes. A concentration 6.25 microM BL-I and 12.5 microM ROS blocked over 93.3 +/- 2.5% of oocytes in germinal vesicle (GV) stage during a 24-hr culture period. Following a second 24-hr culture step in maturation medium (IVM) almost all (91.5 +/- 3.0%) inhibited oocytes resumed meiosis and reached the metaphase II (MII) stage. The MII kinetics was different for inhibited and control oocytes. Fifty percent MII was reached at 13-14 hr in BL-I + ROS treated oocytes, compared to 18 hr in control oocytes. Therefore, control oocytes were fertilised (IVF) after 22 hr IVM and inhibited oocytes after 16 or 22 hr IVM. After IVF, percentage of grade 1 freezable embryos on day 7 (D + 7) as well as percentage of blastocyst formation on D + 8 in the group of BL-I + ROS treated oocytes fertilised after 16 hr IVM were higher (P < 0.05) compared with the other experimental group fertilised after 22 hr IVM but not different in comparison with the control. Survival to freezing and thawing of grade 1 embryos frozen on D + 7 was employed as viability criteria and was similar in all groups. Thus, the presence of BL-I + ROS in the prematuration medium of bovine oocytes determines a reversible meiotic block, without compromising their subsequent developmental competence.     

Resting potential of the mouse neuroblastoma cells: I. The presence of K+ channels activated at high K+ concentration but closed at low K+ concentration including the physiological concentration

The effects of changes in extracellular K⁺ concentration ([K⁺]_o) on the resting membrane potential, the input resistance and ⁸⁶Rb efflux (as a marker of K⁺ efflux) were examined with use of the cultured mouse neuroblastoma cells (N-18 clone). The results obtained are as follows. (1) The membrane potential was depolarized, with an increase in [K⁺]_o at concentrations above 10–20 mM at a rate of 55–58 mV per 10-fold change in [K⁺]_o, but practically unchanged with varying [K⁺]_o below this concentration. (2) Above the critical [K⁺]_o of 10–20 mM, the input membrane resistance decreased sharply by a factor of $\text{1}\text{4}\text{?}\text{1}\text{5}$ with an increase in [K⁺]_o. A similar decrease in the resistance occurred even under the conditions that the membrane potential was held at control level (about ?55 mV) by a steady-state current passage. (3) Elimination of Na⁺ and Cl^? from the external solution brought about practically no change in the membrane potential. (4) A fractional escape rate of ⁸⁶Rb from N-18 cells remained constant at relatively low level (0.125%/min on average) in the low [K⁺]_o range, but increased sharply with increasing [K⁺]_o above 15 mM (e.g., approx. 3.4- and 4.5-fold at 30 and 100 mM [K⁺]_o, respectively). (5) The high K⁺-induced ⁸⁶Rb efflux was not practically inhibited by 1 mM tetraethylammonium or 0.1 mM 4-aminopyridine, indicating that the K⁺ channels activated by an elevation of [K⁺]_o are not the delayed (voltage-dependent) K⁺ channels. The present results favoured the conclusion that N-18 cells carry K⁺ channels which open at high [K⁺]_o but are closed at low [K⁺]_o including the physiological range for the mouse neuroblastoma cells (around 5.4 mM). This conclusion leads to the notion that in the mouse neuroblastoma N-18 cells the K⁺ permeability does not mainly contribute to determining the resting membrane potential under physiological conditions.     

Multiple modes of calcium-induced calcium release in sympathetic neurons I: attenuation of endoplasmic reticulum Ca2+ accumulation at low [Ca2+](i) during weak depolarization.

Many cells express ryanodine receptors (RyRs) whose activation is thought to amplify depolarization-evoked elevations in cytoplasmic Ca2+ concentration [Ca2+](i) through a process of Ca2+ -induced Ca2+ release (CICR). In neurons, it is usually assumed that CICR triggers net Ca2+ release from an ER Ca2+ store. However, since net ER Ca 2+ transport depends on the relative rates of Ca2+ uptake and release via distinct pathways, weak activation of a CICR pathway during periods of ER Ca accumulation would have a totally different effect: attenuation of Ca2+ accumulation. Stronger CICR activation at higher [Ca2+](i) could further attenuate Ca2+ accumulation or trigger net Ca2+ release, depending on the quantitative properties of the underlying Ca2+ transporters. This and the companion study (Hongpaisan, J., N.B. Pivovarova, S.L. Colgrove, R.D. Leapman, and D.D. Friel, and S.B. Andrews. 2001. J. Gen. Physiol. 118:101-112) investigate which of these CICR "modes" operate during depolarization-induced Ca2+ entry in sympathetic neurons. The present study focuses on small [Ca2+](i) elevations (less than approximately 350 nM) evoked by weak depolarization. The following two approaches were used: (1) Ca2+ fluxes were estimated from simultaneous measurements of [Ca2+](i) and I(Ca) in fura-2-loaded cells (perforated patch conditions), and (2) total ER Ca concentrations ([Ca](ER)) were measured using X-ray microanalysis. Flux analysis revealed triggered net Ca2+ release during depolarization in the presence but not the absence of caffeine, and [Ca2+](i) responses were accelerated by SERCA inhibitors, implicating ER Ca2+ accumulation, which was confirmed by direct [Ca](ER) measurements. Ryanodine abolished caffeine-induced CICR and enhanced depolarization-induced ER Ca2+ accumulation, indicating that activation of the CICR pathway normally attenuates ER Ca2+ accumulation, which is a novel mechanism for accelerating evoked [Ca2+](i) responses. Theory shows how such a low gain mode of CICR can operate during weak stimulation and switch to net Ca2+ release at high [Ca2+](i), a transition demonstrated in the companion study. These results emphasize the importance of the relative rates of Ca2+ uptake and release in defining ER contributions to depolarization-induced Ca2+ signals.     

Effect of N-ethylmaleimide on leucine transport in chang liver cells. I. Stimulatory effect on Na+-independent transport at low concentrations of leucine

Pretreatment of Chang liver cells with $\text{N-}\text{ethylmaleimide}$ (0.5 or 1 mM) stimulated Na⁺-independent uptake of leucine at low concentrations (?1 mM). The stimulatory effect of $\text{N-}\text{ethylmaleimide}$ on the uptake of leucine measured in Na⁺-replete medium was completely blocked by the addition of $\text{b-2-}\text{aminobicyclo}\text{[2,2,1]}\text{heptane}\text{-2-}\text{carboxylate}$ (5 mM), which shows that the L system participates in the stimulation. The Na⁺-dependent uptake of glycine was depressed by $\text{N-}\text{ethylmaleimide}$ pretreatment. The stimulation of the Na⁺-independent component of leucine uptake continued for at least 30 min after $\text{N-}\text{ethylmaleimide}$ treatment, while the inhibition of glycine uptake was progressive with time and the Na⁺-dependent uptake of leucine became depressed later, after the treatment. It has been demonstrated that treatment of cells with $\text{N-}\text{ethylmaleimide}$ is capable of increasing the Na⁺-independent influx of leucine and at the same time slightly decreasing the efflux of it. These results suggest that $\text{N-}\text{ethylmaleimide}$ attacks the Na⁺-independent system of amino acid transport at the reactive SH groups(s) of relevant protein(s) in favor of specific activation of that system in this cell.