Mechanochemical study of conformational transitions and melting of Li-, Na-, K-, and CsDNA fibers in ethanol–water solutions

Highly oriented fibers of Li-, Na-, K-, and CsDNA were prepared with a previously developed wet spinning method. The procedure gave a large number of equivalent fiber bundle samples (reference length, L₀, typically = 12–15 cm) for systematic measurements of the fiber length L in ethanol–water solutions, using a simple mechanochemical set up. The decrease in relative length L/L₀ with increasing ethanol concentration at room temperature gave evidence for the B-A transition centered at 76% (v/v) ethanol for NaDNA fibers and at 80 and 84% ethanol for K- and CsDNA fibers. A smaller decrease in L/L₀ of LiDNA fibers was attributed to the B-C transition centered at 80% ethanol. In a second type of experiment with DNA fibers in ethanol–water solutions, the heat-induced helix–coil transition, or melting, revealed itself in a marked contraction of the DNA fibers. The melting temperature T_m, decreased linearly with increasing ethanol concentration for fibers in the B-DNA ethanol concentration region. In the B-A transition region, Na- and KDNA fibers showed a local maximum in T_m. On further increase of the ethanol concentration, the A-DXA region followed with an even steeper linear decrease in T_m. The dependence on the identity of the counterion is discussed with reference to the model for groove binding of cations in B-DNA developed by Skuratovskii and co-workers and to the results from Raman studies of the interhelical bonds in A-DNA performed by Lindsay and co-workers. An attempt to apply the theory of Chogovadze and Frank-Kamenetskii on DNA melting in the B-A transition region to the curves failed. However, for Na- and KDNA the T_m dependence in and around the A-B transition region could be expressed as a weighted mean value of T_m of A- and B-DNA. On further increase of the ethanol concentration, above 84% ethanol for LiDNA and above about 90% ethanol for Na-, K-, and CsDNA, a drastic change occurred. T_m increased and a few percentages higher ethanol concentrations were found to stabilize the DNA fibers so that they did not melt at all, not even at the upper temperature limit of the experiments (～ 80°C). This is interpreted as being due to the strong aggregation induced by these high ethanol concentrations and to the formation of P-DNA. Many features of the results are compatible with the counterion–water affinity model. In another series of measurements, T_m of DNA fibers in 75% ethanol was measured at various salt concentrations. No salt effect was observed (with the exception of LiDNA at low salt concentrations). This result is supported by calculations within the Poisson–Boltzmann cylindrical cell model. © 1994 John Wiley & Sons, Inc.     

Vascular smooth muscle contractile properties in the turtle Pseudemys scripta elegans

1. Turtle aortic rings were characterized by high frequency spontaneous contractile activity and variable responsiveness to constrictor agents.2. The tissue response was remarkably insensitive to temperature at a range of 37°–15°C.3. The contractile response was effectively blocked by the calcium channel antagonist nifedipine and was substantially dependent on extracellular calcium concentrations.4. Lowering the sodium concentration of the bath medium resulted in a strong, transient contraction followed by reduced responsiveness to norepinephrine and the absence of spontaneous activity.5. Disruption of the vessel endothelium resulted in enhanced and reduced responsiveness to norepinephrine (NE) and acetylcholine (ACh), respectively.6. The results indicate that the regulation of contractile function in turtle vascular smooth muscle differs in several respects from that of mammalian tissue, perhaps, reflecting the adaptation of the vasculature to low pressure and ectothermic conditions.     

Regional dependency of the vascular smooth muscle cell contribution to the mechanical properties of the pig ascending aortic tissue

BackgroundDilation and dissection of aneurysmal ascending aortic tissues occur preferentially at the outer curvature of the vessel. In this study we hypothesize that the density and contractile properties of the vascular smooth muscle cells (VSMCs) of the pig ascending aorta (AA) are heterogeneous and could explain the non-uniform remodeling and weakening of the AA during aneurysm formation.MethodsEleven pig AA rings were collected. Two square samples of 15×15 mm were taken from each ring from the inner and outer curvature of the AA. Each sample was subjected to equi-biaxial tensile testing in Krebs–Ringer solution maintained at 37 °C. Each test consisted of 8 cycles of preconditioning followed by one experimental run from 0% to 30% strain. Phenylephrine (10^?5 M) was added to contract VSMCs. After biaxial testing, samples were paraffin-embedded and stained with hematoxylin–phloxine–saffron (HPS) to quantify VSMC density.ResultsSignificant differences in cell density, maximum contractile stress resultant magnitude (MCSRM) and orientation (θ_MCSR) were found between the inner and outer curvature. The inner curvature had the greatest contraction. The outer curvature had the highest VSMC density with the maximum contraction stress resultant oriented towards the axial direction.ConclusionVSMC activation with phenylephrine had a significant effect on the stiffness of the pig AA. This effect was independent of location and direction. However, cell orientation, density and contractile properties were dependent on location and suggest variations in the remodeling capabilities, tissue strain and cell phenotype between locations.     

Studies with dextran 40 in cryopreservation of blood

Whole blood from healthy donors was washed twice with phosphate-buffered saline (PBS) and then resuspended in sufficient PBS to give a final concentration of 2 × 10⁹/cells/ml. Aliquots were combined with equal volumes of the required diluents to give final dextran 40 concentrations of 0, 5, 10, 15, and 20% in PBS. Fifty-lambda samples in 50-lambda Micropets (Clay Adams) were frozen in alcohol baths at temperatures ranging from ?10 to ?80 °C. The specimens were frozen either for 1 min or 16 min, rapidly thawed, and resuspended in PBS or PBS plus dextran. Percentage of hemolysis was determined colorimetrically. Results indicate that concentraitons as low as 5% dextran exert a cryoprotective effect. Increased dextran concentration increases cryoprotection at high subzero bath temperatures (?10 ° and ?20 °C). Dextran concentrations beyond 12% have a damaging effect at low subzero bath temperatures (below ?30 °C). Based on this a two-factor hypothesis for cryopreservation is proposed. Apparent partial recovery of red blood cells without dextran or with 5% dextran during subzero storage was demonstrated.     

Conformation of LiDNA in solutions of LiCl

We have derived radii of gyratin, R_g, from the absolute intensity of the scattered light of mondisperse linear Col E₁ LiDNA in solution at various LiCl concentrations up to 5M. The second virial coefficients, A₂, decrease strongly with increasing LiCl concentration, and vanish between 3 and 5M LiCl. It was thus possible to calculate a limiting value at a high salt concentration of 28.5 nm for the persistence length, a₀, of LiDNA, without the necessity of applying excluded-volume corrections. The value obtained is in good agreement with the value previously obtained for NaDNA at high NaCl concentrations, and can be identified with the high salt limit of DNA flexibility, with long-range electrostatic interactions effectively screened. Sedimentation coefficients in the ultracentrifuge and apparent and translational diffusion coefficients (at finite and vanishing scattering vectors, respectively) from dynamic laser-light scattering have also been obtained up to 5M LiCl. From the sedimentation and apparent diffusion, D(90), (at 90° scattering angle only) above 5M, and up to 9M LiCl, it could be shown that the solutions are stable for reasonable periods of time, and the molecular parameters vary smoothly and moderately at high salt. Conformational transitions were not observed and precipitation occurs between 9 and 10M LiCl.     

Investigation of molecular motion in collagen using the spin-probe technique.

The spin-probe technique was employed to study molecular motion in collagen and modified collagen samples in the ?160° to +200°C region. The effect of water content in the 0–30-wt. % range, relative to vacuum dried samples, on the electron spin resonance spectrum of the probe was also investigated. The spectra at the lowest temperatures consisted of a broad asymmetric triplet. Narrowing of this triplet above ?40° to ?70°C and the appearance of additional lines in the spectrum, interpreted as due to a narrow triplet, at a temperature dependent on the water content were observed. For samples with low water contents [(0–0.4)%] the broad triplet was that expected for a glassy system up to 150–190°C; for these samples the narrow triplet appears at a temperature above 50°C, its intensity increasing with increasing temperature up to 70–100°C, then decreasing with a further temperature increase. For samples with water contents near 30%, the narrow triplet first completely appears at about 0°C, and reaches relative intensities of 35% at 30°C. The motion taking place in collagen and related samples is discussed in terms of these results.     

Temperature dependence of partial volumes of proteins

The change of the apparent partial specific volumes of egg albumin, bovine serum albumin, bovine methemoglobin, β-lactoglobulin, and lysozyme with temperature through the thermal transitions of the proteins have been measured with dilatometers. Four regions in the plot of the apparent partial specific volumes against temperature can be recognized: (1) linear sections extending from 25°C up to 45–50°C: (2) a decrease in slope between 50°C and 60°C; (3) a sharp increase in slope with increasing temperature coinciding with the appearance of heat coagulation of the protein and followed by (4) a decrease in the slope. The return of the protein samples to 25°C yields linear relations between the apparent partial specific volumes of the heat-denatured proteins and the decreasing temperature.     

A continuum model of contraction waves and protoplasm streaming in strands of Physarum plasmodium

We present a mathematical model for continuously distributed mechanochemical autooscillations (autowaves) in a protoplasmic strand of Physarum polycephalum. The model is based on a hypothesis of local positive feedback between deformation and contraction of the contractile apparatus. This feedback is mediated through a cell regulatory system whose kinetics involves coupling to mechanical strain. Mathematical analysis and computer simulations have demonstrated that the solutions of the model agree quantitatively with the available experimental data. In particular, hydrodynamic interaction alone, between different sections of the strand via the streaming endoplasm, is capable of inducing the characteristic contractile behavior.     

Pressure effects on thermal preference behaviour in gammarid amphipods from 600–1000m in Lake Baikal

Temperature preference behaviour of gammarid crustaceans from depths between 600–2000 m in Lake Baikal was studied in a system which provided a stable temperature gradient at pressures ranging from 50–150 atm. At the pressure of their habitat, these animals show well-defined modal distributions of sojourn temperatures around mean values from 3.0–5.5°C, av. 3.9 ± 0.3°C; mean modal T_p is estimated at 3.5°C. Year-round habitat temperatures are 3.0–3.6°C. The effect of changing pressure upon sojourn temperatures was explored over the range 50–150 atm. The slope of the mean sojourn temperature/pressure curves was 2.1°C/100 atm, significantly greater than 0. Mean nodal temperature estimates indicate that the corresponding slope in the range of 50–100 atm is 3°C/100 atm, and in the range of 100–150 atm, is likely to exceed 5°C/100 atm.     

Effect of preservation of human semen sample at 4–6 and 25 °C on sperm motility

Sperm motility is the result of transverse movements that exist along its tail. It plays an important role in male fertility. The aim of this study was to evaluate the influence of keeping washed normozoospermic semen samples at 4–6 and 25 °C on the motility of spermatozoa. 26 semen samples of normozoospermic were washed twice in modified Ham’s F10 medium. Then, thirteen of the semen samples were kept in refrigerator (4–6 °C) and the remaining samples were stored in incubator (25 °C) for 12 days. On the 0 (immediately after sampling as control group), 1st, 2nd, 5th, 7th and the 12th days, the percentage of fast progressive (grade a), slow progressive (grade b), non-progressive (grade c) and immotile (grade d) sperm cells were calculated for each temperature. The data obtained from this study showed that the percentages of a, b and c grades of motile spermatozoa were significantly decreased (p?<?0.001) during 12 days at the both temperatures but reduction of these percentages has a gentle slope at 4–6 °C. There was no motile sperm after 12 days of storage. This study suggests that motile spermatozoa could be retrieved up to 7 days after the storage of washed normozoospermic men semen samples at 4–6 and 25 °C. Also, there were no motile sperm cells 12 days after sampling.     

Cryopreservation of the common carotid artery of the rabbit: Optimization of dimethyl sulfoxide concentration and cooling rate

This paper describes the continuation of studies that demonstrated the suitability of CP-Tes solution as a medium for the introduction and removal of dimethyl sulfoxide in rabbit common carotid arteries and established the kinetics of cryoprotectant permeation in that tissue. In this paper we report the tolerance of rabbit common carotid artery to dimethyl sulfoxide, in concentrations up to 30% (w/w), using a technique of exposure that was designed to control osmotic stress. The maximum concentration achieved without damage was 15% (w/w). Vessels were then equilibrated with 15% dimethyl sulfoxide and cooled to −80 °C at 0.22, 0.69, 2.15, or 9.63 °C/min: they were then transferred to the gas phase of a liquid nitrogen refrigerator {temperature below −160 °C) for storage. Thawing was carried out in a 37 °C water bath. The optimum rate of cooling for these conditions was found to be 0.69 °C/min. The maximal recovery of contractile force in response to 10⁻⁶ M norepinephrine was 30–40%; relaxation to acetylcholine (an endothelium-mediated function) was 80% of control, and an estimated 71% of endothelial cells survived with minimal ultrastructural change.     

The Effect of Nacl on Campylobacter Jejuni/Coli

The growth of 2 strains of Campylobacter jejuni/coli was investigated in 0–2.0 % NaCl in Brucella broth at 35° G and 30° C. Both strains tolerated more NaCl in the growth medium at 35° C than at 30° C. 2 % NaCl was bacteriocidic at both temperatures. The strains also grew in the medium without added NaCl. At 35° C, low concentrations of NaCl stimulated the growth of strain 5616, but not the growth of strain B33. At 30° C, strain 5616 grew in NaCl concentrations up to 1.0 % and strain B33 in 0 % and at the control concentration (0.5 % NaCl). The survival of 22 C. jejuni/coli strains in 2.0 % NaCl at 4° C and 35° C was also investigated. Human strains showed significantly greater tolerance to 2.0 % NaCl at both temperatures than did the strains isolated from animals. These findings suggest that the salting of food can be effective in preventing the growth or survival of C. jejuni/coli.     

Expression of Hsp27 correlated with rat detrusor contraction after acute urinary retention

Heat shock protein 27 (Hsp27) can regulate actin cytoskeleton dynamics and contractile protein activation. This study investigates whether Hsp27 expression is related to bladder contractile dysfunction after acute urinary retention (AUR). Female rats were randomized either to AUR by urethral ligation or to normal control group. Bladder and smooth muscle strip contraction at time points from 0 h to 7 days after AUR were estimated by cystometric and organ bath studies. Hsp27 expression in bladder tissue at each time point was detected with immunofluorescence, Western blots, and real-time PCR. Expression of the three phosphorylated forms of Hsp27 was detected by Western blots. Smooth muscle ultrastructure was observed by transmission electron microscopy. Data suggest that maximum detrusor pressure and both carbachol-induced and spontaneous detrusor strip contraction amplitude decreased gradually for the duration from 0 to 6 h, and then increased gradually to near-normal values at 24 h. Treatment of muscle strips with the p38MAK inhibitor, SB203580, inhibited carbachol-induced contractions. Smooth muscle ultrastructure damage was the highest at 6 h after AUR, and then lessened gradually during next 7 days, and ultrastructure was close to normal. Expressions of Hsp27 mRNA and protein and the proteins of the three phosphorylated forms were higher at 0 h, decreased to lower levels up to 6 h, and then gradually increased. Therefore, we conclude that rat bladder contractile function after AUR worsens during 0–6 h, and then gradually recovers. The findings of the current study suggest that Hsp27 modulates bladder smooth muscle contraction after AUR, and that phosphorylation of Hsp27 may be an important pathway modulating actin cytoskeleton dynamics in bladder smooth muscle contraction and reconstruction after injury.     

Smooth muscle contraction: mechanochemical formulation for homogeneous finite strains

Chemical kinetics of smooth muscle contraction affect mechanical properties of organs that function under finite strains. In an effort to gain further insight into organ physiology, we formulate a mechanochemical finite strain model by considering the interaction between mechanical and biochemical components of cell function during activation. We propose a new constitutive framework and use a mechanochemical device that consists of two parallel elements: (i) spring for the cell stiffness; (ii) contractile element for the sarcomere. We use a multiplicative decomposition of cell elongation into filament contraction and cross-bridge deformation, and suggest that the free energy be a function of stretches, four variables (free unphosphorylated myosin, phosphorylated cross-bridges, phosphorylated and dephosphorylated cross-bridges attached to actin), chemical state variable driven by Ca2+-concentration, and temperature. The derived constitutive laws are thermodynamically consistent. Assuming isothermal conditions, we specialize the mechanical phase such that we recover the linear model of Yang et al. [2003a. The myogenic response in isolated rat cerebrovascular arteries: smooth muscle cell. Med. Eng. Phys. 25, 691-709]. The chemical phase is also specialized so that the linearized chemical evolution law leads to the four-state model of Hai and Murphy [1988. Cross-bridge phosphorylation and regulation of latch state in smooth muscle. Am. J. Physiol. 254, C99-C106]. One numerical example shows typical mechanochemical effects and the efficiency of the proposed approach. We discuss related parameter identification, and illustrate the dependence of muscle contraction (Ca2+-concentration) on active stress and related stretch. Mechanochemical models of this kind serve the mathematical basis for analyzing coupled processes such as the dependency of tissue properties on the chemical kinetics of smooth muscle.     

Raman spectroscopy of the milk globule membrane and triglycerides

The acyl chain mobilities of the lipids of bovine milk fat globules and the component triglycerides have been determined by Raman spectroscopy as a function of temperature from -100°C to 80°C. A broad transition is observed centered about 2–6°C. The C-H and C-C stretching bands in the spectra of liquid and crystalline triglycerides are used comparatively to show that the lipids of the milk globule membrane are 30–40% more ordered than the lipids of the intact milk fat globules at 20°C. Synthetic triglyceride melts, quenched rapidly, are used to illustrate the effect of the thermal history of a sample on lipid order as determined spectroscopically.Strong infrared amide I and amide II bands at 1646 and 1543 cm^?1, respectively, indicate that the protein conformation of the globule membrane is not characterized by extensive regions of beta-sheet structure. Raman spectra of the globule triglycerides indicate cis unsaturation of $\text{39 ± 5\%}$ by comparison to triolein and trielaidin.     

Effects of temperature and molecular interactions on the vibrational infrared spectra of phospholipid vesicles

Infrared spectra were obtained as a function of temperature for a variety of phospholipid/water bilayer assemblies (80% water by weight) in the 3000-950 cm^?1 region. Spectral band-maximum frequency parameters were defined for the 2900 cm^?1 hydrocarbon chain methylene symmetric and asymmetric stretching vibrations. Temperature shifts for these band-maximum frequencies provided convenient probes for monitoring the phase transition behavior of both multilamellar liposomes and small diameter single-shell vesiclesof dipalmitoyl phosphatidylcholine/water dispersions. As examples of the effects of bilayer lipid/cholesterol/water (3 : 1 mol ratio) and lipid/cholesterol/amphotericin B/water (3 : 1 : 0.1 mol ratios) vesicles were examined using the methylene stretching frequency indices. In comparison to the pure vesicle form, the transition width of the lipid/cholesterol system increased by nearly a factor of two (to 8°C) while the phase transition temperature remained approximately the same (41° C). For the lipid/cholesterol/amphotericin B system, the phase transition temperature increased by about 4.5° C (to 45.5°C) with the transition width increasing by nearly a factor of four ($\text{to}\text{≈ 15°}\text{C}$) above that of the pure vesicles. The lipid/cholesterol/amphotericin B data were interpreted as reflecting the formation below 38°C of a cholesterol/amphotericin B complex whose dissociation at higher temperature (38–60°C range) significantly broades the gel-liquid crystalline phase transition.     

Effects of temperature on photosynthesis and growth of seagrasses

The effect of temperature on the photosynthesis and growth of seagrasses may be summarized by considering the ways in which temperature alters the characteristics of the photosynthesis-irradiance (P-I) curve of seagrasses. Within the limits of physiological tolerance (∼6–30°C) temperature has little effect on the initial slope of the P-I curve. At 35–40°C the photosynthetic capacity of seagrasses is reduced. Within the limits of physiological tolerance, the rate of photosynthesis at light saturation, the dark respiration rate and the light compensation point more than double as temperature increases. The optimum temperature for photosynthesis decreases from 25–35°C at light saturation to as low as 5°C as irradiance decreases. As a result of these effects of temperature on the P-I curve, growth of seagrasses in high(saturating) light environments increases with temperature, whereas growth of seagrasses in low (near the light compensation point) light environments decreases as temperature increases.     

The effects of aqueous neutral-salt solutions on the melting temperatures of deoxyribonucleic acids

The helical stability of a variety of DNA samples, ranging in base composition from 0 to 72 mole-% GC, has been studied by heat denaturation at neutral pH in increasing concentrations of LiCl, NaCl, KCl, CsCl, Li₂SO₄, and K₂SO₄. The variation of melting temperature with average base composition, dT_m/dX_GC, was found to decrease drastically in the concentrated salt media, e.g., from 41°C in 0.006M LiCl to 29°C in 3.2M LiCl, and from 39°C in 0.003M Li₂SO₄ to 18°C in 1.6M Li₂SO₄. At the same time, the thermal transition is much more cooperative in the concentrated salt solutions than at low ionic strength. Indeed, at limiting salt concentrations, the transition breadth seems to reach a minimum value irrespective of the compositional heterogeneity of the DNA samples. Attempts to correlate the observed decrease of dT_m/dX_GC with predicted changes in the enthalpy of melting, deduced from a simple theoretical treatment, experimental data on the binding of counterions and water to DNA, and experimental data on thermal denaturation, were unsuccessful. However, the strongly reduced composition dependence of the melting temperature can be understood in terms of a destabilizing effect of the concentrated salt media on GC-base pairs. It is suggested, though not proven, that the destabilization involves the displacement of water molecules from the DNA helix.