One-electron Reduction of 8-hydroxy-5-deazaisoalloxazine in Aqueous Solution: A Pulse Radiolysis Study

Summary

The one-electron reduction of 8-hydroxy-5-deazaisoalloxazine (HMDI) has been studied in aqueous solution in the acidity range pH0 to 13 using the reducing species CO₂·^?, e^?_aq and (CH₃)₂COH radicals. The spectral and other properties of the HMDI radicals were found to be independent of the reductant used. Four protolytic forms of the radical were distinguished with associated pK_a values of 2·3 ± 0·3, 6·0 ± 0·3 and 10·1 ± 0·3.     

Causation of Cancer. British Medical Bulletin,Vol. 14, No. 2

Summary

Peroxyl radicals of poly(U), poly(A), and single- and double-stranded DNA have been produced by photolysing H₂O₂ in oxygenated aqueous solution in presence of the substrates. The peroxyl radicals are formed by the reaction of OH radicals with the polynucleotides followed by addition of oxygen.

The lifetime of the peroxyl radicals and the rate constant of their reactions with the thiols cysteamine, gluthathione and dithiothreithol have been measured by time-resolved e.s.r. spectroscopy. The unusually long lifetimes range from 0·2 to 3·3 s. The activation energy for the decay for all four substrates is 10·3 ± 1 kcal/mol (43 kJ mol^?1). The reaction rate constants with the thiols range from k = 0·8 × 10⁴ to 1·3 × 10⁵ dm³ mol^?1 s^?1. The reactions of the thiols with the peroxyl radical of poly(U) are known to prevent strand break formation. This shows that the peroxyl radicals of poly(U) observed by e.s.r. are intermediates in the pathway leading to strand break formation.     

Pulse Radiolytic Study of the Interaction of SO−·4 with Deoxynucleosides. Possible Implications for Direct Energy Deposition

Summary

Using the technique of pulse radiolysis it has been demonstrated that the interaction of SO^?·₄ with deoxynucleosides (k ～ < 2 × 10⁸–2·3 × 10⁹ dm³ mol^?1 s^?1) in aqueous solution at pH 7·0 results in the formation of the corresponding one-electron oxidized radicals which either deprotonate or hydrate to yield OH adducts. Based upon the ease of oxidation of the deoxynucleosides, dG, dA, dC, dT, by SO^?·₄, the apparent redox potentials are in the order dG ? dA ≈ dC > dT. With the exception of deoxyuridine, the deoxynucleoside radicals produced on interaction with SO^?·₄ have been shown to have oxidizing properties based upon the interactions with tetranitromethane and the nitroxyls, TMPN and NPPN. The deoxynucleoside radicals (dG, dA and dC) do not interact with oxygen (k < 10⁶ dm³ mol^?1 s^?1) in contrast to the interaction observed with the thymidine radical (k = 2·5 × 10⁷ dm³ mol^?1 s^?1). The implications of these findings are presented in terms of the properties of the discussed radicals as relating to those of potential DNA base radicals (positive centres) produced by direct energy deposition within DNA. The use of SO^?·₄ to mimic, to some extent, the effects of direct energy deposition in DNA may assist in our understanding of the resulting molecular processes relevant to radiobiological studies.     

Subject Index

Summary

Using conductivity detection, pulse radiolysis experiments showed that solvent protonation of the electron adducts of cytosine, 5-methyl cytosine and 2′-deoxycytidine occurs with rate constants k ≥ 2 × 10⁴M^?1s^?1. The protonated electron adducts transfer an electron to p-nitroactetophenone (PNAP) with rate constants ranging from 3·5×10⁹ to 5·3·10⁹M^?1s^?1. The transfer is quantitative (G = 2·7), as shown by conductometric and spectroscopic measurements. In the presence of O₂ no electron transfer to O₂ takes place, implying that O₂ adds to the protonated electron adduct radicals.

No electron transfer from the H- and OH-adducts of the cytosine derivatives, either to PNAP or to O₂, takes place near neutral pH. It is suggested that the differences in the reaction behaviour of the H-adduct radicals and the protonated electron adduct radicals towards PNAP can be accounted for if different radicals are formed by H-addition and protonation of the electron adduct. The H atoms most probably add to the C-5-C-6 double bonds, whereas the electron adducts are protonated at N-3 and/or 0-2.     

X-ray Induced Free Radicals in Single Crystals of N-acetyl-L-leucine

Summary

Double-stranded DNA from calf thymus was irradiated in the presence of bovine serum albumin (BSA) at a ratio of 1:10 by mass under air, N₂ or N₂O, with and without ethanol, tert-butanol or isopropanol in 10^?2 mol dm^?3 sodium phosphate buffer, pH 7·0. The irradiated solutions were analysed by a nitrocellulose filter assay and by high-performance liquid gel chromatography. The results show that OH^· radicals as well as hydrated electrons induce protein—DNA crosslinks. Further, alcohol radicals react with BSA producing protein radicals which become crosslinked to DNA. If OH radicals are scavenged by alcohol, strand breakage of DNA does not occur.     

The Reactions of the Hydroxymethyl Radical with 1,3-dimethyluracil and 1,3-dimethylthymine

Summary

Hydroxymethyl radicals ·CH₂OH, generated by the radiolysis of methanol (0·5 mol dm^?3) in N₂O-saturated aqueous solutions, were reacted with 1,3-dimethyluracil or 1,3-dimethylthymine (10^?3) mol dm^?3). The products were identified and their G values determined. It has been concluded that in 1,3-dimethyluracil ·CH₂OH attack occurs only at C(6) while in 1,3-dimethylthymine there is partitioning between addition (two-thirds) and H-abstraction from the C(5)-methyl group (one-third). A rate constant for CH₂OH addition to 1,3-dimethyluracil of about 10⁴ dm³ mol^?1 s^?1 is estimated. Complexities that may arise in the radiolysis of pyrimidines such as 1,3-dimethylthymine, apparently as a consequence of the formation of 5-alkylidenepyrimidines, are discussed. A value of 0·15 has been estimated for the disproportionation/combination ratio for the hydroxymethyl radical self-termination reaction.     

Rates for Repair of pBR 322 DNA Radicals by Thiols as Measured by the Gas Explosion Technique: Evidence that Counter-ion Condensation and Co-ion Depletion are Significant at Physiological Ionic Strength

Summary

Rates of repair of pBR 322 plasmid DNA radicals by thiols of varying net charge (Z) at pH 7 and physiological ionic strength were measured using the oxygen explosion technique. The extent of conversion of supercoiled to relaxed circular plasmid was measured by HPLC as a function of the time of oxygen exposure before or after irradiation, the time-courses being fitted by a pseudo-first-order kinetic expression with k₁ = k₂[RSH]. Values of k₂ (M^?1 s^?1) were: 2·1 × 10⁵ (GSH, Z = ?1), 1·4 × 10⁶ (2-mercaptoethanol, Z = 0), 1·2 × 10⁷ (cysteamine, Z = +1), 6·6 × 10⁷ (WR-1065 or N-(2-mercaptoethyl)-1,3-diaminopropane, Z = +2). The approximately 6-fold increase in rate with each unit increase in Z is attributed to concentration of cationic thiols near DNA as a consequence of counter-ion condensation and reduced levels of anionic thiols near DNA owing to co-ion depletion. The results are quantitatively consistent with chemical repair as a significant mechanism for radioprotection of cells by neutral and cationic thiols under aerobic conditions, but indicate that repair by GSH will compete effectively with oxygen only at low oxygen tension.     

The efficacy of weekly and bi-weekly heat training to maintain the physiological benefits of heat acclimation

ObjectivesTo examine the efficacy of weekly and bi-weekly heat training to maintain heat acclimatization (HAz) and heat acclimation (HA) for 8 weeks in aerobically trained athletes.DesignRandomized, between-group.MethodsTwenty-four males (mean [m ± standard deviation [sd]; (age, 34 ± 12 y; body mass, 72.6 ± 8.8 kg, VO_2peak, 57.7 ± 6.8 mL·kg^?1·min^?1) completed five trials (baseline, following HAz, following HA (HAz + HA), four weeks into heat training [HT_WK4], and eight weeks into HT [HT_WK8] that involved 60 min of steady-state exercise (59.1 ± 1.8% vVO_2peak) in an environmental laboratory (wet bulb globe temperature [WBGT], 29.6 ± 1.4 °C) on a motorized treadmill. Throughout exercise, heart rate (HR) and rectal temperature (T_rec) were recorded. Following HAz + HA, participants were assigned to three groups: control group (HT₀), once per week heat training (HT₁), and twice per week heat training (HT₂). HT involved heated exercise (WBGT, 33.3 ± 1.3 °C) to achieve hyperthermia (38.5–39.75 °C) for 60 min. Repeated measures ANOVAs were used to determine differences.ResultsHAz + HA resulted in significant improvements in HR (p < 0.001) and T_rec (p < 0.001). At HT_WK8, HR was significantly higher in HT₀ (174 ± 22 beats?min^?1) compared to HT₂ (151 ± 17 beats?min^?1, p < 0.023), but was not different than HT₁ (159 ± 17 beats?min^?1, p = 0.112). There was no difference in % change of T_rec from post-HAz + HA to HT_WK4 (0.6 ± 1.3%; p = 0.218), however, HT_WK8 (1.8 ± 1.4%) was significantly greater than post-HAz + HA in HT₀ (p = 0.009).ConclusionsBi-weekly HT provided clear evidence for the ability to maintain physiological adaptions for 8 weeks following HA.     

Lung Tumour Induction in Mice after X-rays and Neutrons

Summary

Dose–response curves were determined for pulmonary adenomas and adenocarcinomas in mice after single acute doses of 200 kVp X-rays and cyclotron neutrons (E¯ = 7·5 MeV). A serial-killing experiment established that the radiation induction of chromosome aberrations. The validity of the concept of PLD neous cancers. The dose versus incidence (I) of tumours in male and female mice for X-ray doses between 0·25 and 7·5 Gy is ‘bell-shaped’ and best fitted with a purely quadratic induction and exponential inactivation terms, i.e. I = A + BD²e^?αD. In contrast, the tumour dose–response after 0·1–4·0 Gy of neutrons is best fitted by I = A + BD e^?αD and is steeply linear ≤ 1 Gy, peaks between 1 and 3 Gy and sharply declines at 4·0 Gy. The data for the female mice ≤ 1 Gy neutrons are best fitted to the square root of the dose.

A major objective of the experiments was to derive neutron RBE values. Because of the differences between the X-ray (quadratic) and neutron (linear) curves, the RBE_n will vary inversely with decreasing X-ray dose. The RBE values at 1 Gy of X-rays derived from the B coefficients in the above equations are 7·4 ± 3·2 (male and female); 8·6 ± 3·6 (female) and 4·7 ± 1·8 (male). These are high values and imply even higher values at the doses of interest to radiation protection. If, however, one restricts the analysis to the initial, induction side of the response (≤ 1 Gy neutrons, ≤ 3 Gy X-rays) then good linear fits are obtainable for both radiations and indicate neutron RBE values of 7·4 ± 2·3 for female mice and 4·5 ± 1·8 for males, and these are independent of dose level.     

Radicals Produced by the Reactions of SO−4 with Uridine and Its Derivatives. Studies by Pulse Radiolysis and γ-radiolysis

Summary

Using a pulse radiolysis technique, the sulphate radical, SO^?₄, has been demonstrated to react with uridine, 3′-UMP and 5′–UMP to produce intermediates of a new type, decaying according to a first-order rate law with k = 2·1 4·1, 3·0 × 10⁵ s^?1 respectively at pH7. The rate for uridine increases slightly with increasing pH. With deoxyuridine and 2′, 3′-isopropylidene uridine, on the other hand, OH adduct radicals with longer lives (second-order decay) were found in accordance with the known fact that similar radicals are produced from N(1)-methylated uracils with SO^?₄. On γ-irradiation of similar solutions containing the substrates that produce such short-lived species, chain reactions and high yields of unaltered uracil were found. Some optical properties are detailed and a probable structure of the short-lived species is discussed in relation to the substituent effect at the 2′ position of the sugar part and to its strong reducing ability revealed by the rapid reaction with TNM.     

The influence of the b‐value combination on apparent diffusion coefficient based differentiation between malignant and benign tissue in cervical cancer

Purpose:

To analyze the influence of different b‐value combinations on apparent diffusion coefficient (ADC)‐based differentiation of known malignant and benign tissue in cervical cancer patients.

Materials and Methods:

A total of 35 patients with stage IB1, IB2, IIA cervical cancer underwent a 3.0T MRI scan prior to radical hysterectomy and pelvic lymph node dissection. Conventional T1‐ and T2‐weighted sequences and a diffusion‐weighted sequence (b = 0, 150, 500, 1000 seconds/mm²) were performed. Regions‐of‐interest (ROI) were drawn on ADC maps derived from five different b‐value combinations (0, 500; 0, 150, 500; 0, 1000; 0, 150, 500, 1000; 150, 500, 1000 seconds/mm²). The influence of the b‐value combination on ADC‐based differentiation of benign and malignant tissue was analyzed using receiver‐operating‐characteristics curves.

Results:

For all b‐value combinations, ADCs were significantly lower (P < 0.001) in cervical malignancies (1.15 ± 0.21·10^?3; 1.10 ± 0.21·10^?3; 0.97 ± 0.18·10^?3; 0.97 ± 0.23·10^?3 and 0.85 ± 0.18·10^?3 mm²/second respectively to the aforementioned b‐value combinations) than in benign cervix (2.08 ± 0.31·10^?3; 2.00 ± 0.29·10^?3; 1.62 ± 0.23·10^?3; 1.54 ± 0.21·10^?3 and 1.42 ± 0.22·10^?3 mm²/second respectively). The diagnostic accuracy was high for all b‐value combinations and without statistical differences between the combinations.

Conclusion:

ADC‐based differentiation of benign from malignant cervical tissue is independent of the tested b‐value combinations. The results support the inclusion and possible pooling of studies using different b‐value combinations in meta‐analyses on ADC‐based tissue differentiation in cervical cancer. J. Magn. Reson. Imaging 2010;32:376–382. © 2010 Wiley‐Liss, Inc.

     

Stem Cells in Bone and Bone Marrow after Contamination with Bone-seeking Radionuclides

Summary

The effects of time, mass and oxidation state on plutonium gastrointestinal absorption and tooth adsorption were studied during and after chronic ingestion of plutonium-238 (IV) or (VI) (1·55–15·60 kBq/ml) in 6·5 mm bicarbonate medium by fed rats via drinking water for 8 days to 3 months. Animals were killed during the ingestion to follow the kinetics of whole-body storage and clearance of plutonium. At 1·55 kBq/ml the amount of plutonium retained in the skeleton increased continuously during the 85 days of ingestion and reached a plateau thereafter. This plutonium retention was therefore dependent on the total mass administered but not proportional to this mass, as the fraction of administered plutonium retained decreased during the first 22 days of ingestion and then stabilized. This is reflected by the gastrointestinal transfer (f₁), which had risen to (3·80 ± 0·82) × 10^?5 on Day 3 of ingestion and then decreased to a stabilized value of (1·07 ± 0·06) × 10^?5 from Day 30 to the end of the ingestion period. In the liver, the amount of plutonium retained reached a plateau, which lasted from Day 30 to the end of ingestion. The kidneys and spleen were also found to be retention sites. By Day 3 of ingestion, for a mass ingested of 5 × 10^?7 g/kg of body mass, the maximum mean value of f₁ we found was smaller than the 10^?4 recommended by ICRP Report 30. The oxidation state had no effect on f₁. Large plutonium deposition was observed on the teeth. For both oxidation states (IV) and (VI), about 0·10% of the administered dose was deposited on the teeth after 3 days of ingestion, whatever the plutonium concentration administered. However, whereas the amount of plutonium (IV) deposited did not change throughout the ingestion period, tooth deposition of plutonium (VI) decreased.     

Reactions of OH Radicals with Poly(U) in Deoxygenated Solutions: Sites of OH Radical Attack and the Kinetics of Base Release

Summary

Pulse radiolysis of N₂O-saturated solutions of poly(U) in the presence of tetranitromethane showed that 81 per cent of the radicals formed are reducing in nature. Using data from other sources it has been estimated that 70 per cent of the OH radicals add to the base at C(5) and 23 per cent at C(6) while only 7 per cent abstract an H-atom from the sugar moiety. To a large extent the C(5) OH adduct radicals attack the sugar moiety of poly(U) thereby inducing strand breakage and base release. G (base release) = 2·9 can be subdivided into three components: (a) immediate (20 per cent), (b) fast (50 per cent) and (c) slow (30 per cent). The immediate base release must occur either during the free-radical stage or as a result of the rapid (t_½ < 4 min at 0°C) decomposition of a diamagnetic product. The fast and the slow processes are only readily observable at elevated temperatures, e.g. at 50°C the half lives are 83 min and 26 h, respectively (E_a (fast) = 68 kJ mol^?1, E_a (slow) = 89 kJ mol^?1, A (fast) = 1·5 × 10⁷ s^?1, A (slow) = 1·9 × 10⁹ s^?1. It is concluded that there are three different types of sugar lesions giving rise to base release, structures for which are tentatively proposed.     

Interactions of Thiyl Free Radicals with Oxygen: A Pulse Radiolysis Study

Summary

A pulse radiolysis study of glutathione in aqueous solution at pH 5·5 containing N₂O/O₂ mixtures at various ratios indicates that oxygen rapidly adds to the thiyl glutathione radical yielding a transient absorption, with a maximum at 540 nm, whose characteristics appear to be compatible with assignment to the GSOO^· radical. The reaction appears to be an equilibrium whose kinetic constants have been estimated (k_f = 2·0 × 10⁹ dm³ mol^?1 s^?1, k_b = 6·2 × 10⁵ s^?1). Evidence for electron transfer from ascorbate to the GSOO^· radical has been obtained and the respective rate constant has been determined to be 1·75 ± 0·15 × 10⁸ dm³ mol^?1 s^?1.     

Model Studies for the Direct Effect of High-energy Irradiation on DNA. Mechanism of Strand Break Formation Induced by Laser Photoionization of Poly U in Aqueous Solution

Summary

Laser flash photolysis of polyuridylic acid (poly U) in anoxic aqueous solutions leads to biphotonic photoionization of the uracil moiety followed by the formation of single strand breaks (ssb). The rate constant for ssb formation (1·0 s^?1, obtained from the slow component of conductivity increase at 23°C and pH 6·8) increases with decreasing pH to 235 s^?1 at pH 3·5. The activation energy (pre-exponential factor) was measured to be 66 kJ mol^?1 (5 × 10¹¹ s^?1) at pH 6·8. Addition of dithiothreitol (DTT) or glutathione (GSH) prevents ssb formation by reacting with a poly U intermediate (rate constant = 1·2 × 10⁶ and 0·16 × 10⁶ dm³ mol^?1 s^?1, respectively). Since with OH radicals as initiators very similar data have been obtained for the kinetics of ssb formation and for the reaction with DTT, we conclude that photoionization of the uracil moiety in poly U leads eventually to the same chemical pathway for ssb formation as that induced by OH radicals. Furthermore, we propose that protection by DTT and GSH occurs via H donation to the C-4′ radicals of the sugar moiety of DNA and to the C-4′ and the C-2′ radicals of poly U.     

Additivity of Type O and Type N Damage in Diploid Yeast

Summary

Idiopathic pneumonitis is a major cause of morbidity and mortality in patients with leukaemia undergoing total body irradiation (TBI) and bone marrow transplantation (BMT). The effect of variation in dose rate of TBI on the development of lethal and sublethal lung damage has been investigated in mice by measuring changes in carbon monoxide uptake. CBA mice were irradiated using a ⁶⁰Co source at 0·02, 0·05, 0·1, 0·2, 0·5 and 1·0 Gy min^?1 to a total dose of 15·5 Gy. A log–linear relationship between the severity of impairment of carbon monoxide uptake (V_CO) and dose rate was found. Ventilatory requirement (ventilation rate/V_CO) was raised 20 to 40 weeks after TBI at dose rates above 0·1 Gy min^?1. Time of onset and extent of elevation of ventilatory requirement were also dose-rate dependent. The implications of these findings for clinical practice are discussed.     

Association for Radiation Research/Netherlands Radiobiological Society

Summary

The kinetics of pulse radiolytically induced intramolecular radical transformations: Trp/N^·→Tyr/O^·, Tyr/O^·→Trp/N^·, Met/S∴Br→Trp/N^· and Met/S∴Br→Tyr/O^· has been investigated at various pH levels and temperatures in model peptides: Trp-(Pro)_n-Tyr, Trp-(Gly)_n-Tyr, Trp-(Pro)_n-Met (n = 0–3), Tyr-Phe-Met-Arg-Phe-NH₂·2AcOH, Met⁵-enkephaline and [d-Ala]²Met⁵-enkephaline. The rate constants of the Trp/N^·→Tyr/O^· transformation at pH 8 were found to decrease exponentially with the distance between Trp and Tyr in the proline peptides, while in the glycine peptides the rate of the transformation is less dependent on the number of bridging glycine residues. The activation energies determined fall into the range 10–20 kJ mol^?1 irrespective of: (i) the ionization state of tryptophyl radical and tyrosine, (ii) type of bridging amino acids, and (iii) reversal of the direction of the electron transfer upon tyrosine OH group ionization. The activation entropies at 298 K for the peptides of the glycine and proline series are negative and rather high, and fall into the relatively narrow range of ?90 to ?140 J mol^?1 deg^?1. These activation parameters seem to indicate that a tunnelling mechanism is involved in the electron transfer between strictly oriented aromatic moieties of Trp and Tyr. The variation of the activation parameters with average separation distance in the case of Trp-(Pro)_n-Tyr shows a predominance of the electronic factor over the nuclear in determining the distance dependence of the electron transfer rate. The intramolecular Met/S∴Br→Tyr/O^· transfer proceeds with the rate constant of 1·1 × 10⁵ s^?1 in Met⁵-enkephaline and 5·7 × 10⁴ s^?1 in [d-Ala]²Met⁵-enkephaline. The activation parameters for this transformation E_a = 30 kJ mol^?1 and ΔS⁴₂₉₈ = ?62 J mol^?1 deg^?1 are close to those of the Trp/N^·→Tyr/O^· transformation, suggesting a similar mechanism for the electron transfer.     

Chemical Properties of Water-soluble Porphyrins. 5. Reactions of Some Manganese (III) Porphyrins with the Superoxide and Other Reducing Radicals

Summary

Solution properties of three manganese porphyrins, in monomeric form, were investigated. These were the ‘picket-fence-like’ porphyrin Mn(III)-α,α,α,β-tetra-ortho(N-methylisonicotinamidophenyl)porphyrin (Mn(III)PFP) and two ‘planar unhindered’ porphyrins, the Mn(III)TMPyP (tetrakis (4-N-methylpyridyl)porphyrin) and Mn(III)TAP (tetra(4-NNN-trimethyl-anilinium)porphyrin). The porphyrin properties studied were: the absorption spectra in their manganic and manganous forms; acid/base properties of the aquo complexes; the effect of potential axial ligands (up to a concentration of 0·1 mol dm^?3) and their one electron reduction potentials. Knowing these properties, the reaction of the Mn(III) porphyrins with the superoxide radical and other reducing radicals were studied using the pulse radiolysis technique. The second-order reaction rate constant of O^?₂ with the Mn(III) porphyrins, which governs the catalytic efficiency of the metalloporphyrins upon the disproportionation of the superoxide radical, was 5·1 × 10⁷ to 4·0 × 10⁵ dm³ mol^?1s^?1, depending on the pH and the nature of the metalloporphyrin. These values are at least one order of magnitude lower than found for Fe(III)TMPyP. One electron reduction of the three Mn(III) porphyrins by e^?_aq, CO^?₂, CH₂OH and (CH₃)₂COH had similar second-order rate constants (10⁹-10¹⁰ dm³ mol^?1 s^?1). That for (CH₃)₂(CH₂)COH was about 10⁵ dm³ mol^?1 s^?1. Reduction in all cases produced the corresponding Mn(II) porphyrin and no intermediate was found. The oxidation reaction of the Mn(II) porphyrins by O^?₂ was approximately two orders of magnitude faster when compared to the reduction of Mn(III) porphyrins with the same radical. Since the reactivities of O^?₂ towards the three manganese (III) compounds follow their reduction potentials, it is suggested that these reactions are governed by an outer-sphere mechanism. This suggestion is corroborated by the finding that water molecules acting as axial ligands, in these aqueous solution systems, are not replaced by another potential ligand when the latter is in the concentration range of 100 mm or less.     

Long-term Changes in Mouse Lung Following Inhalation of a Fibrosis-inducing Dose of 239PuO2: Changes in Collagen Synthesis and Degradation Rates

Summary

Mice were exposed by nose-only inhalation to ²³⁹PuO₂, which resulted in an IAD of 1110 ± 29 Bq. At various times after exposure, rates of collagen metabolism were measured using validated in vivo methods based on the administration of radiolabelled proline, together with a large flooding dose of unlabelled proline and measurement of its incorporation into lung collagen as hydroxyproline. Dramatic increases in both synthesis and degradation rates of collagen were observed. At 54 days after exposure the fractional synthesis rates in experimental mice were almost five times those in controls (control: 3·2 ± 0·6%/day, ²³⁹PuO₂-exposed: 14·5 ± 0·4%/day) and by 300 days synthesis rates, although declining, were still more than double the control values. A similar pattern of change was observed for collagen degradation. The combination of changes in synthesis and degradation rates led to a 60% increase in lung collagen content by 300 days (control: 3·05 ± 0·24 mg/lung, ²³⁹PuO₂-exposed: 4·88 ± 0·42 mg/lung). The data suggest that extensive remodelling of the lung connective tissue matrix occurs during development of fibrosis and that, over long periods of time, small imbalances between synthesis and degradation may result in quite large increases in protein content.     

Comparison of different test protocols to determine maximal lactate steady state intensity in swimming

ObjectivesThis study compared step test, lactate minimum (LM) test and reverse lactate threshold (RLT) test protocols with maximal lactate steady state (MLSS) in free-swimming. All test protocols used fixed duration increments and high work-rate resolution (≤ 0.03 m·s^?1) to ensure high sensitivity.DesignValidation study.Methods23 swimmers or triathletes (12 male and 11 female) of different ages (19.0 ± 5.9 yrs) and performance levels (400 m personal best 1.38 ± 0.13 m·s^?1, FINA points 490 ± 118) completed an incremental step test (+0.03 m·s^?1 every 3 min) to determine speed at 4 mmol·L^?1 and at modified maximal distance method, a LM test, a RLT test and two to five 30 min tests (±0.015 m·s^?1) to determine MLSS. Following a 200 m all-out and a 5 min rest, LM was determined during an incremental segment (+0.03 m·s^?1 every 2 min) as the nadir of the speed-lactate curve. After a priming segment with four increments (+0.06 m·s^?1), RLT was determined as the lactate apex during a reverse segment (?0.03 m·s^?1) every 3 min.ResultsThe mean differences (± limits of agreement) to speed at MLSS were +1.0 ± 4.1% (speed at 4 mmol·L^?1), +1.5 ± 3.5% (modified maximum distance method), ?0.2 ± 4.7% (LM) and 2.0 ± 3.1% (RLT). All threshold concepts showed good agreement with MLSS pace (intraclass correlation coefficient ≥ 0.886).ConclusionsTest protocols with a fixed step duration and fine increments allowed high accuracy in estimating MLSS pace. With similar criterion agreement to the LM and RLT tests, incremental step tests appear more practicable due to less prior knowledge required and derivation of individual training zones.