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PubMed1989

Dual regulatory mechanisms of proton transport in rat papillary collecting duct cells in culture.

The regulation of proton transport and cytosolic pH was studied in rat papillary collecting duct (PCD) cells in culture using a pH-sensitive fluorescence probe, 2,7-bis-carboxyethyl-5,6-carboxyfluorescein (BCECF). Data were obtained from confluent monolayers grown on glass coverslips and dipped in a HCO3- -free medium, pH 7.40. The resting intracellular pH (pHi) was 7.16 +/- 0.03 (n = 20). When PCD cells had been acidified by pretreatment with NH4Cl, pHi immediately recovered toward the resting value. Two mechanisms participated in this recovery: a Na+-dependent mechanism which could be inhibited by amiloride (indicative of Na+-H+ exchanger) and a Na+-independent process (a proton ATPase). The pHi recovery from acid loading was inhibited by amiloride to about 55% of the control recovery (half-maximal effect at 100 microM). The rate of pHi recovery after the readdition of Na+ to a sodium-free medium exhibited saturation kinetics (half maximal rate at 28 mM). Dicyclohexylcarbodiimide (DCCD), an inhibitor of a plasma membrane proton ATPase, and the depletion of cellular ATP induced by 2 mM potassium cyanide (KCN) also partially inhibited the rate of pHi recovery after cell acidification with a NH4Cl load. When PCD cells were treated with 1 mM DCCD, amiloride almost completely inhibited pHi recovery. Amiloride and the removal of external Na+ had induced a gradual fall in pHi to a new resting value and rapidly recovered when Na+ was added. We conclude that PCD cells grown in culture have at least two proton transport mechanisms: a Na+-H+ exchanger and a plasma membrane proton ATPase. The kinetics of these processes can be reliably assessed by the pH-sensitive fluorescent probe, BCECF. Both the Na+-H+ exchanger and the plasma membrane proton ATPase may contribute to urinary acidification.

Adenosine Triphosphatases↗
PubMed2001

Discrimination of brain abscess and cystic tumor by in vivo proton magnetic resonance spectroscopy.

Proton magnetic resonance (MR) spectroscopy was evaluated for the differentiation of brain abscesses and cystic brain tumors. Proton MR spectroscopy was performed in vivo in two patients with brain abscess and eight patients with various cystic brain tumors (anaplastic astrocytoma, glioblastoma, and metastatic brain tumor). MR imaging with contrast medium demonstrated ring-like enhanced mass lesions in all patients. The various resonance peaks in proton MR spectra were assigned to metabolites according to chemical shifts. Treatment of the cystic brain lesions was based on the information from proton MR spectroscopy. Aspirated pus from one patient with brain abscess was examined using ex vivo proton MR spectroscopy. The in vivo spectra of brain abscess contained resonance peaks attributed to acetate, lactate, alanine, amino acids, and lipids in both cases, and an additional peak of succinate in one case. In vivo spectra of the neoplasms contained resonance peaks corresponding to lactate, lipids, choline, creatine, and N-acetyl aspartate. Proton MR spectroscopy is useful for discriminating brain abscess from cystic tumors with similar neuroimaging appearance, which is very important for determining the treatment strategy.

Acetates↗
PubMed1991

The narrow proton beam therapy unit at the the Svedberg Laboratory in Uppsala.

The synchrocyclotron at the The Svedberg Laboratory (TSL) in Uppsala is now reconstructed and can presently operate with fixed frequency and proton energies up to 100 MeV. A first treatment room with a narrow proton beam unit for therapy of eye tumours is now in operation. Therapy of eye melanomas started in April, 1989 and during 1989 and 1990, 19 patients were treated with 72 MeV protons. The narrow beam unit provides a fixed horizontal beam and the patient is treated in a seated position. The present paper describes mainly the technical aspects of the unit which so far has been used only for eye melanomas. In the future, modifications of the unit will allow therapy of intracranial targets when higher proton energies are available. In its final form, the proton therapy facility at TSL will harbour a second treatment unit. Here a rotating gantry for 200 MeV protons will provide a broad beam, which will enable treatment of tumours located anywhere in the body.

Eye Neoplasms↗
PubMed2005

Charged particle equilibrium of small field in clinical proton beams.

It is expected that proton beam radiotherapy will become an effective treatment for tumors. For an organ for which a correct dose prescription is required, a proton beam has the ability to provide the dose most suitable for the specific purpose. We measured the charged particle equilibrium factor of proton beams in water using a plane parallel ionization chamber. The maximum energy of the proton beam used in this study was 70 MeV, produced from an isochronous cyclotron. We assume that the charged particle equilibrium factor can be separated into longitudinal and lateral components, that is, the factor E (Z, R) is dependent on depth, Z, and field radius, R; such that E (Z, R) =E (Z) E (R). The E (Z) -factor of primary protons was considered in order to investigate the influence of secondary charged particles. From the results, the charged particle equilibrium factor for the longitudinal component does not remain sharp with the decrease of water depth, and the lateral component is not maintained with the decrease of field size. However, the longitudinal components of primary protons at shallow depth were in equilibrium.

Protons↗
PubMedDate not supplied

[Electron and proton transport in chloroplasts taking into account lateral heterogeneity of thylakoids. Mathematical model].

A mathematical model of a chloroplast was constructed, which takes into account the inhomogeneous distribution of complexes of photosystems I and II between granal and intergranal thylakoids. The structural and functional complexes of photosystems I and II, which are localized in intergranal and granal thylakoids, respectively, and the b/f complex, which is uniformly distributed in thylakoid membranes, are assumed to be immobile. The interactions between spatially distant electron transport complexes are provided by plastoquinone and plastocyanine, which diffuse in the thylakoid membrane and intrathylakoid space, respectively. The main stages of proton transport associated with the functioning of photosystem II and oxidation-reduction transformations of plastoquinone are considered. The model takes into account the interactions of protons with membrane-bound buffer groups, the lateral diffusion of hydrogen ions in the intrathylakoid space and in the lumen between adjacent granal thylakoids, and the transmembrane proton transport associated with the function of ATP synthase and passive leakage of protons from thylakoids outside. The numerical integration of two systems of differential equations describing the behavior of some variables in two different regions: granal and intergranal thylakoids was performed. The model describes adequately the kinetics of processes being studied and predicts the occurrence of inhomogeneous lateral profiles of proton potentials and redox state of electron carriers. Modeling the electron and proton transport with allowance for the topological features of chloroplasts (lateral heterogeneity of thylakoids) is important for correct interpretation of "power-flux" interactions and the experimentally measured kinetic parameters averaged over the entire spatially inhomogeneous thylakoid system.

Biological Transport↗
PubMed2001

New techniques in hadrontherapy: intensity modulated proton beams.

Inverse planning and intensity modulated (IM) X-ray beam treatment techniques can achieve significant improvements in dose distributions comparable to those obtained with forward planned proton beams. However, intensity modulation can also be applied to proton beams and further optimization in dose distribution can reasonably be expected. A comparative planning exercise between IM X-rays and IM proton beams was carried out on two different tumor cases: a pediatric rhabdomyosarcoma and a prostate cancer. Both IM X-rays and IM protons achieved equally homogenous coverage of the target volume in the two tumor sites. Predicted NTCPs were equally low for both treatment techniques. Nevertheless, a reduced low-to-medium dose to the organs at risk and a lesser integral non-target mean dose for IM protons in the two cases favored the use of IM proton beams.

Dose-Response Relationship, Radiation↗
PubMed2003

Long-term effects of low-dose proton radiation on immunity in mice: shielded vs. unshielded.

BACKGROUND: Outside the protection of the terrestrial environment, astronauts on any long-term missions will unavoidably be exposed to fields of charged particle radiation dominated by protons. These fields and their biological risks are modified in complex ways by the presence of protective shielding. METHODS: To examine the long-term effects of space-like proton exposures on immune status, we treated female C57BL/6 mice with 3 or 4 Gy of 250 MeV monoenergetic protons or the complex space-like radiation field produced after 250 MeV protons are transported through 15 g x cm(-2) aluminum shielding. The animals were euthanized 122 d post-irradiation and lymphocyte phenotypes, hematological parameters, and lymphocyte blastogenesis were characterized. RESULTS: There were significant dose-dependent decreases in macrophage, CD3+/CD8+ T, NK, platelet, and red blood cell populations, as well as low hematocrit and hemoglobin levels. In contrast, dose-dependent increases in spontaneous, but not mitogen-induced, blastogenesis were noted. The differences in dose composition between pristine and shielded proton fields did not lead to significant effects in most measures, but did result in significant changes in monocyte and macrophage populations and spontaneous blastogenesis in the spleen. CONCLUSIONS: The data indicate that whole body exposure to proton radiation at doses of the order of large solar particle events or clinical treatment fractions may have long-term effects on immune system status.

Animals↗
PubMed2003

Low energy proton irradiation effects on alveolar macrophages from young and aged rats.

A prospective study was undertaken to assess the radiotoxicity of accelerated particles in pulmonary alveolar macrophages (AM). We evaluated the effects of a single dose (10-75 Gy) of an external low-energy (20 MeV) proton beam on cultured AM oxidative metabolism and phagocytic function. Macrophages are the first line of defense against invading pathogens and are known to generate superoxide anion (O2), nitric oxide (NO), and mediators of antimicrobial and antitumoral defense mechanisms. We obtained AM by bronchoalveolar lavage from young (1-2 month old) and aged (9-12 month old) male Wistar rats. Cell viability, phagocytosis, O2 and NO production in control and proton-irradiated cultured AM were evaluated The effect of proton irradiation on cell viability was dose-dependent The higher doses induced a dramatic decrease in viability in the aged population. Phagocytosis increased 1.3-1.4 fold inboth populations irrespective of the dose delivered. Generation of O2 was always higher in the aged population for all the doses assayed and showed no significant variation from the control values. In the young population a clear increase was observed with doses of 25 and 50 Gy. NO production in AM from young animals rose in a dose-dependent manner. Conversely, proton irradiation did not affect NO production in macrophages from aged animals. The results of this study demonstrate that AM isolated from young and aged rats are functionally different and show a distinct behavior when exposed to proton irradiation. These findings suggest that age may condition response and must be taken into account when accelerated particle-radiotherapy protocols are considered as a valid therapeutic option for the treatment of cancer. To the best of our knowledge, this is the first report comparing sham-irradiated and proton-irradiated young and aged AM.

Aging↗
PubMedDate not supplied

[Neoplastic transformation of mouse fibroblasts under the influence of high-energy protons and gamma-rays].

Oncoginic transformations of mouse fibroblasts C3H10T1/2 after exposure to proton energies 150 and 584 MeV were compared with fibroblast effects of gamma-radiation. Prior to exposure, cell populations (2.7 x 10(3) cells/cm2) were inoculated in plastic vials with the surface area of 75 cm2 and cultivated 11 days. Survivability was determined by comparing the number of cell colonies in irradiated and non-irradiated (control) vials. Transformation rate was calculated by dividing the total transformation focus number by the number of survived cells in a vial. Rate of oncogenic transformations after gamma- and proton (584 MeV) irradiation was essentially identical, i.e. the parameter grew rapidly at the doses < 1 Gy and slowed down at the doses > 1 Gy. In the dose interval between 1 and 5 Gy, transformation rate for proton energy 150 MeV was found low compared with gamma-radiation and proton energy 584 MeV. It is hypothesized that the different transformation rate after exposure to proton energy 150 MeV is linked with the high linear energy transfer as compared with the proton energy of 584 MeV and gamma-radiation.

Animals↗
PubMedDate not supplied

Nature of lysozyme-water interactions by proton NMR.

Proton spin-lattice relaxation measurements were performed in 10 mM lysozyme solution as a function of temperature and degree of substitution of solvent H2O with D2O. The results show that in the temperature range from 274 to 323 K, the intermolecular lysozyme proton water proton coupling contributes appreciably to the observed water proton relaxation rate. In this system exchange between water protons and labile protein protons does not dominate the behaviour with temperature of the water-lysozyme intermolecular contribution to the spin-lattice relaxation.

Kinetics↗
PubMed1991

Biological effectiveness of high-energy protons: target fragmentation.

High-energy protons traversing tissue produce local sources of high-linear-energy-transfer (LET) ions through nuclear fragmentation. We examine the contribution of these target fragments to the biological effectiveness of high-energy protons using the cellular track model. The effects of secondary ions are treated in terms of the production collision density using energy-dependent parameters from a high-energy fragmentation model. Calculations for mammalian cell cultures show that at high dose, at which intertrack effects become important, protons deliver damage similar to that produced by gamma rays, and with fragmentation the relative biological effectiveness (RBE) of protons increases moderately from unity. At low dose, where sublethal damage is unimportant, the contribution from target fragments dominates, causing the proton effectiveness to be very different from that of gamma rays with a strongly fluence-dependent RBE. At high energies, the nuclear fragmentation cross sections become independent of energy. This leads to a plateau in the proton single-particle-action cross section, below 1 keV/micron, since the target fragments dominate.

Cell Survival↗
PubMed1990

Functional reassembly of the coated vesicle proton pump.

We have shown previously that treatment of the coated vesicle proton-translocating adenosine triphosphatase (H(+)-ATPase) with chaotropic agents results in the release of a set of peripheral polypeptides which includes the 73-, 58-, 40-, 34-, and 33-kDa subunits (Adachi, I., Puopolo, K., Marquez-Sterling, N., Arai, H., and Forgac, M. (1990) J. Biol. Chem. 265, 967-973), with a coordinate loss of H(+)-ATPase activity. In the present paper we report the functional reassembly of the coated vesicle proton pump following dissociation of the peripheral subunits. Reassembly was demonstrated by restoration of ATP-driven proton transport using both native membranes and reconstituted vesicles and by Western blot analysis using a monoclonal antibody specific for the 73-kDa subunit. Reassembly occurs by attachment of a peripheral subcomplex containing the 73-, 58-, 34-, and 33-kDa subunits together with the 40-kDa polypeptide. The reassembled H(+)-ATPase, like the native proton pump, is inhibited by N-ethylmaleimide, 7-chloro-4-nitrobenz-2-oxa-1,3-diazole, and N,N'-dicyclohexylcarbodiimide. Reassociation shows a biphasic time dependence, with restoration of 50-60% of the starting proton transport activity in the 1st h followed by recovery of a further 20-30% of the activity after 24 h. Reassembly also shows a marked dependence on protein concentration but, unlike solubilization of the intact H(+)-ATPase complex, does not require the presence of glycerol. Despite the ability of nucleotides to promote dissociation of the peripheral complex by chaotropic agents, reassociation is not blocked by the presence of 1 mM ATP. These results thus provide the first evidence for functional reassembly of a vacuolar H(+)-ATPase complex and should be useful in further analysis of the role of individual subunits in the assembly and activity of these ATP-driven proton pumps.

Adenosine Triphosphate↗
PubMed1975

Relationship of the light-induced proton uptake in bovine retinal outer segment fragments to triton-induced membrane disruption and to volume changes.

Light-induced proton uptake in bovine retinal outer segment (ROS) fragments was shown to be closely related to pH, salt concentration, membrane integrity, and perhaps secondarily to the volume of osmotic compartments. The principal findings were as follows: 1. As pH increased, both the discs and the plasmalemma swelled, and proton uptake markedly diminished. 2. As the discs were disrupted by increasing concentrations of Triton, proton uptake at slightly alkaline pH was supplanted by proton release. 3. Increasing the concentration of chloride salts caused increased H+ uptake roughly proportional to osmotic shrinkage of the ROS. Buffering by acetate prevented the measurement of proton uptake in the presence of acetate salts, although osmotic behavior of the ROS was similar to that observed in chloride salts. Although increasing the concentration of sucrose also resulted in osmotic shrinkage of the ROS, it was not accompanied by a systematic increase in the magnitude of proton uptake. 4. Light-induced H+ uptake was accompanied by small but reproducible changes in volume, probably of the discs. The magnitude and direction of these rapid volume changes were subject to influence by pH, solute, and other variables.

Animals↗
PubMedDate not supplied

Acute skin reactions observed in fractionated proton irradiation.

Between May 1985 and July 1987, 49 skin reactions of 43 patients treated by proton irradiation were observed at the Particle Radiation Medical Science Center (PARMS), The University of Tsukuba. Taking the peak skin score as an endpoint, the radiobiological effects [relative biological effectiveness (RBE) and time-dose relationships] of the proton beam in multi-fractionated treatments were estimated. Factors influencing the skin dose, such as the prescribed tumor dose, tumor site, and number of applied fields, were also analyzed. The following conclusions regarding acute skin reactions to the clinical use of proton irradiation were obtained: 1) the physical skin-sparing effect of proton irradiation in single-field irradiation, especially in superficial regions, is not large compared with that of high-energy photon irradiation; 2) multidirectional proton irradiation significantly reduced the skin dose and severity of acute reactions; 3) the radiobiological effects of the proton beam, RBE and the time factor, estimated in human skin in multi-fractional treatment were slightly smaller than those of X-rays, i.e., 0.92 and -0.25 +/- 0.09, respectively.

Dose-Response Relationship, Radiation↗
PubMed1988

Effect of an uncE ribosome-binding site mutation on the synthesis and assembly of the Escherichia coli proton-translocating ATPase.

Plasmid pRPG54, which carries the genes for the eight subunits of the proton-translocating ATPase of Escherichia coli, has been found to carry a single base change of a G to an A in the ribosome-binding site for uncE, the gene which codes for the N,N'-dicyclohexylcarbodiimide-binding subunit c of the Fo. This noncoding region mutation both lowers expression of uncE by a factor of 2-3 and affects the function of the ATPase, specifically of the Fo sector. The presence of the mutation results in a decrease in the proton permeability of the Fo or of the entire F1Fo-ATPase complex when either is synthesized from genes on a multicopy plasmid. Expression of uncE from an F1Fo plasmid carrying the wild type ribosome binding site results in increased membrane proton permeability and decreased ability of the resultant ATPase to couple a transmembrane proton gradient to ATP synthesis both in vitro and in vivo. Also, although an Fo plasmid carrying the correct ribosome-binding site causes harmful, F1-dependent proton permeability in unc+ cells (Brusilow, W. S. S. (1987) J. Bacteriol. 169, 4984-4990), an identical plasmid carrying the mutation does not, even though it still codes for a functional reconstitutable Fo. The results show a relationship between the relative level of expression of uncE from a multicopy plasmid and the assembly pathway, proton permeability, and energy-coupling characteristics of the ATPase.

Adenosine Triphosphate↗
PubMed1985

Proton translocation by a native and subunit III-depleted cytochrome c oxidase reconstituted into phospholipid vesicles. Use of fluorescein-phosphatidylethanolamine as an intravesicular pH indicator.

The existence of a proton pump associated with bovine cytochrome c oxidase (EC 1.9.3.1) has over the last few years been a matter of considerable dispute. In an attempt to resolve some of the problems with the measuring system we have synthesized fluorescein-phosphatidylethanolamine which when reconstituted with cytochrome c oxidase into phospholipid vesicles provided a reliable indicator of the intravesicular pH. It was observed that cytochrome c oxidase catalyzed the abstraction of almost 2 protons from the intravesicular medium/molecule of ferrocytochrome c oxidized. In parallel experiments whereby the extravesicular pH was measured with an electrode it was found that the enzyme appeared to be responsible for the appearance of almost 1.0 proton/molecule of ferrocytochrome c oxidized. Taken together these data unequivocally demonstrate that cytochrome c oxidase behaves as a proton pump. Furthermore, the other proton which was abstracted is believed to be used for the process of the reduction of oxygen. Similar experiments were performed with a cytochrome c oxidase preparation which was devoid of subunit III. Under these circumstances the enzyme appeared to be unable to translocate protons across the vesicular membrane but was competent to abstract protons from the intravesicular medium for the reduction of oxygen.

Animals↗
PubMed1985

Proton therapy at Harvard.

Fractionated precision high-dose proton radiotherapy has been carried out at the Harvard Cyclotron Laboratory (HCL) since 1973, in a collaborative effort with the Radiation Medicine Department of Massachusetts General Hospital (MGH) and the Retina Service of the Massachusetts Eye and Ear Infirmary (MEEI). This paper will discuss proton treatment in general, treatment planning procedures, and results to date in major patient categories. 846 patients have been treated with fractionated proton therapy at the Harvard Cyclotron, with normal tissue and tumor responses consistent with an RBE of 1.1 for the proton beam. Proton beam therapy is the treatment of choice for patients with uveal melanomas, and chordomas and chondrosarcomas involving the skull base and cervical spine. Improved dose distribution possible with protons have allowed greater doses than are given conventionally to be delivered to patients with prostatic carcinoma, head and neck malignancies, ano-rectal cancers, and retroperitoneal tumors. Doses employed have been usually 10 to 20% greater than normally would be delivered in our department to such tumors. Generally, local control rates have been good.

Animals↗
PubMed1986

Proton translocation by cytochrome oxidase vesicles catalyzing the peroxidatic oxidation of ferrocytochrome c.

Coupled with the peroxidatic oxidation of ferrocytochrome c under anaerobic conditions, proteoliposomes reconstituted with a purified preparation of bovine heart cytochrome oxidase ejected protons into the external medium with an apparent H+/e- ratio of 0.9. At the same time, protons in the intravesicular space were consumed. Dicyclohexylcarbodiimide significantly inhibited the proton translocation. Cyanide (0.14 mM) completely inhibited both the peroxidase and proton translocating activities. On the contrary, in the presence of 1 mM CO the proton ejection was abolished almost completely, but 50% of the peroxidase activity persisted. This result suggests the operation of multiple mechanisms in the peroxidase reaction and that the CO-sensitive one is coupled to the proton translocation.

Animals↗