Tumour growth modifies intravascular polyamine transport by plasma lipoproteins in the mouse.

Polyamines are polycationic compounds which are implicated in cell division and tumor growth. We have evaluated the potential role of plasma lipoproteins in the transport of major polyamines, spermine, spermidine and putrescine, and the effect of tumor growth on such transport. Plasmas of healthy male BL6/DBA2 mice and of mice bearing Lewis lung carcinoma (3LL) were fractionated by isopycnic density gradient ultracentrifugation, and polyamine content determined in lipoprotein fractions. Spermidine was the most abundant polyamine in the lipoproteins of both control and tumor-bearing mice and was principally associated with HDL (d: 1.046-1.136 g/ml); approx. 40% of total plasma polyamines was lipoprotein-associated in control mice and 60% in cancerous mice. Only minor amounts were transported by LDL (< 10% of total lipoprotein-associated polyamines), while VLDL were devoid of these substances. Marked elevations of circulating levels of LDL were found in 3LL grafted mice: in these particles however, the contents of spermidine and spermine were significantly reduced. A preferential uptake of polyamines by red blood cells could in part explain this marked reduction of LDL polyamine content, but the consequence of this reduction on the net electrical charge and biochemical function of LDL remains unknown. Elevations of plasma LDL and HDL levels in 3LL-grafted mice underlie the finding that only minor modification was detected in the putrescine content of these particles. However, it is evident that elevated total amounts of putrescine were present in the plasma of such animals. Finally, the density profile of polyamines was modified in cancerous mice in which a shift to transport in lighter apo.AI-containing HDL particles was observed for spermidine; an even more marked shift was found for spermine. In conclusion, our data demonstrate that HDL particles constitute the major plasma vehicle for polyamine transport in both control and in tumor-bearing mice.

Is biomedical nuclear magnetic resonance limited by a revisitable paradigm in physics?

The history of nuclear magnetic resonance (NMR) can be divided generally into two phases: before the Second World War, molecular beam methods made it possible to detect the whole set of spins. However, these methods were destructive for the sample and had a very low precision. The publications of F. Bloch and E. Purcell in 1946 opened up a second phase for NMR with the study of condensed matter, but at the expense of an enormous loss in theoretical sensitivity. During more than half a century, the method of Bloch and Purcell, based on inductive detection of the NMR signal, has allowed many developments in biomedicine. But, curiously, this severely constraining limitation on sensitivity has not been called into question during this half-century, as if the pioneers of the pre-war period had been forgotten.

Proton nuclear magnetic resonance study of the binary complex of cytochrome P450cam and putidaredoxin: interaction and electron transfer rate analysis.

A 1H nuclear magnetic resonance study of the complex of cytochrome P450cam-putidaredoxin has been performed. Isocyanide is bound to cytochrome P450cam in order to increase the stability of the protein both in the reduced and the oxidized state. Diprotein complex formation was detected through variation of the heme methyl proton resonances which have been assigned in the two redox states. The electron transfer rate at equilibrium was determinated by magnetization transfer experiments. The observed rate of oxidation of reduced cytochrome P450 by the oxidized putidaredoxin is 27 (+/- 7) per s.

Static and magic angle spinning (31)P NMR spectroscopy of two natural plasma membranes.

Static and magic angle spinning (31)P NMR spectroscopy was used for the first time in natural plasma membranes from erythrocytes and skeletal muscle to study phospholipid arrangement and composition. Typical static powder-like spectra were obtained showing that phospholipids were in a bilayer arrangement. Magic angle spinning narrowed spectra into two components. The first one corresponded to phosphatidylcholine and the second one to the other phospholipids with intensities in agreement with the known phospholipid composition. These findings show that NMR data previously acquired using model membranes can be transposed to studies on phospholipids in their natural environment.

In vivo 31P NMR assessed effects of dantrolene on mechanics and energy metabolism in tetanic stimulated rat gastrocnemius.

Dantrolene does not affect fatigue from submaximal effort and MVC while it decreases twitch tension. We hypothesize that dantrolene could modify the relation between energy metabolism and fatigue by inhibiting calcium release from the sarcoplasmic reticulum. The effects of dantrolene (10 mg) on mechanical and metabolic parameters of gastrocnemius muscle were examined by 31P NMR during an in vivo fatigue test. The fatigue test constituted of three successive 20 min periods of increased stimulation rhythms and followed by a 20 min recovery period. 31P NMR was used to determine phosphocreatine (PCr), ATP and intracellular pH changes, while tension was recorded. We showed that dantrolene increased mechanical fatigue while PCr levels were similar to those from control animals. Acidosis was most prominent in dantrolene treated rats. These results suggest that dantrolene firstly affects calcium cycling with additive effects to fatigue and, secondly, modifies the activation of oxidative metabolism and the energy cost of the generated tension.

Proton NMR relaxation times of experimental Lewis lung carcinoma after irradiation.

NMR proton spin-lattice (T1) and spin-spin (T2) relaxation times were measured ex vivo on Lewis lung carcinoma after in vivo single irradiation with an absorbed dose of 4 Gy. The results were compared to tumoural volume evolution, pathological examinations, and cell kinetic measurements. Tumour growth decreased between the third and the sixth day after irradiation while relaxation times, especially T1, is increased 2 days before the clinical recurrence of the tumour is observed. Pathological morphometric measurements tempted to show that necrosis is less extended after irradiation. Cell cycle analysis demonstrated the G2/M phase blockade by radiation after one day, and its release 4 days later. These phenomena could be important for in vivo radiotherapy follow-up using determination of relaxation times by magnetic resonance imaging (MRI).

Proton magnetic resonance spectroscopy of blood plasma in heart transplant recipients treated with cyclosporine: an early prognosis test of long-term graft tolerance.

BACKGROUND: Previous results have established the potential interest of proton magnetic resonance spectroscopy (MRS) of plasma lipoproteins in the detection of rejection processes after heart transplantation. The aim of this study was to determine whether MRS can provide a relevant long-term prognosis factor as early as 1 week after transplantation. METHODS: Eighteen patients were monitored for a mean period of 16 months after transplantation. The ratio of the sum of the MRS total line widths (TLW) for lipoprotein moieties, obtained 1 week after transplantation and cyclosporine administration, over the same sum obtained on the day of transplantation (TLW(8/0)), as well as the ratio between the corresponding intensities of methyl and methylene moieties (IR) were used to quantify the lipoprotein spectral profile. RESULTS: TLW(8/0), with a cutoff value of 0.8, seemed to have the most value in predicting rejection processes (RP) several months later. All six patients with no RP (good prognosis) and all five patients with three or more RPs (poor prognosis) during the entire 16-month follow-up period were correctly detected as early as 8 days after transplantation. The seven patients with only one or two RPs, mainly occurring during the first months after transplantation, were usually classified by MRS as having good prognosis. CONCLUSIONS: The magnetic resonance spectrum depends on both qualitative and quantitative variations in the different lipoprotein fractions, known to be carriers of cyclosporine. The magnetic resonance spectrum could thus be an early expression of the ability of these lipoproteins to modulate the cyclosporine-mediated immunosuppression.

An in-vivo magnetic resonance imaging study of the olfactory bulbectomized rat model of depression.

The olfactory bulbectomized (OB) rat is a well-accepted animal model of depression. The present magnetic resonance imaging (MRI) investigation demonstrates alterations in signal intensities in cortical, hippocampal, caudate and amygdaloid regions in OB animals, but not in sham operated controls. Ventricular enlargement was also evident in OB animals. These alterations have implications with regard to the face and construct validity of this model.

Time-domain quanification of amplitude, chemical shift, apparent relaxation time T2, and phase by wavelet-transform analysis. Application to biomedical magnetic resonance spectroscopy.

The wavelet-transform method is used to quantify the magnetic resonance spectroscopy (MRS) parameters: chemical shift, apparent relaxation time T2, resonance amplitude, and phase. Wavelet transformation is a time-frequency representation which separates each component from the FID, then successively quantifies it and subtracts it from the raw signal. Two iterative procedures have been developed. They have been combined with a nonlinear regression analysis method and tested on both simulated and real sets of biomedical MRS data selected with respect to the main problems usually encountered in quantifying biomedical MRS, specifically "chemical noise," resulting from overlapping resonances, and baseline distortion. The results indicate that the wavelet-transform method can provide efficient and accurate quantification of MRS data.

In vivo evidence of abnormal mechanical and oxidative functions in the exercised muscle of dystrophic hamsters by 31P-NMR.

Mechanical properties and metabolic adaptation to exercise in skeletal muscle of dystrophic hamsters were studied with an in vivo 31P-NMR multistep fatigue test. Three successive 20-min steps with increasing rhythms of tetanic stimulation were followed by a 20-min recovery period. Fatigue in dystrophic hamsters (DH) developed more rapidly and was greater than in normal hamsters (NH); total mechanical performance per min increased step by step in NH while it decreased in DH, showing a progressive mechanical impairment of the dystrophic muscles. ADP and PCr recovery rates were significantly reduced in DH muscles. Acidosis appeared in both DH and NH and persisted in DH throughout the test, suggesting reduced mitochondrial oxidative capacity of the dystrophic muscle. The pH recovery rate was reduced in DH muscles suggesting a reduction in export protons capacity. These results provide evidence of impaired mitochondrial function and intracellular ionic regulation in the dystrophic muscle, associated with the lack of dystrophin and dystrophin-associated glycoproteins in the DH.

Measurements of the effect of storage at various temperatures on the T1 of ex vivo tissues.

The detailed measurements of the effects of storage at various temperatures on the nuclear magnetic resonance (NMR) spin-lattice relaxation time T1 are reported. On the basis of the results, a method to monitor tissue transformation in hyperthermia of biological systems using magnetic resonance imaging (MRI) techniques is discussed. The method was found on proton relaxation time spin-lattice T1 variation measurements. Parameter F described as tissue structure transformation and/or protein denaturation rate index is discussed.

Effects of dimethylformamide on in vivo fatigue and metabolism in rat skeletal muscle measured by 31P-NMR.

Dimethylformamide (DMF) is widely used as an industrial solvent in spite of well-established hepatotoxicity and adverse effects on in vitro muscle contractility. The doses used in the studies describing these effects were higher than the doses required to solubilize drugs to be injected at very low levels and the potential effects of DMF at very low levels has not yet been explored. The goal of this work was to study the effects of an acute, low dose of DMF (3 mu/100 g body weight, administered i.p.) on mechanical parameters and energy metabolism of contracting rat skeletal muscle. Metabolic changes were followed by 31P nuclear magnetic resonance spectroscopy. Tension was significantly lower during the fatigue test in DMF-treated rats than in controls. Phosphomonoesters and inorganic phosphorus level were lower, and intracellular pH was higher in DMF-treated rats than in controls, showing that energy metabolism was activated to a lesser degree, in relation with the lower mechanical performance, after DMF. Skeletal muscle is a target organ for dimethylformamide which has a major effect on muscle contractility by decreasing the tension developed. The effects of DMF suggest that it is unsuitable for use as a drug vehicle for in vivo injections, even at a very low nonhepatotoxic doses.

Proton nuclear magnetic resonance of regenerating rat liver after partial hepatectomy.

Spin-lattice (T1) and spin-spin (T2) proton nuclear magnetic resonance relaxation times were measured over a 48-hours period of experimental liver regeneration in Wistar rats. T2 showed an early significant increase reaching a plateau 30% above baseline from the 10th hrs onwards. Laparotomized control animals showed no change in T2 values. The increase in T1 occurred at a later stage but was no different from that in laparotomized controls. T1 reached a peak, 20% above baseline, around the 30th hr. The changes observed were far less marked than those previously described for cancer tissue, which showed about a 60% increase in T1. Liver T1 fluctuations followed a circadian pattern, with a minimum at night's end and a maximum around mid-day. No circadian rhythm was seen for T2. The observed T1 and T2 changes were discussed with respect o mitotic and metabolic events known to occur during regeneration of the liver.

High resolution nuclear magnetic resonance spectroscopy in clinical biology: application in oncology.

Nuclear Magnetic Resonance (NMR) technology is mainly known in medicine by the rapidly growing activity in proton imaging (MRI) and, to a lesser extent by the very promising in vivo spectroscopy (MRS). Though about forty years old, the idea of using high resolution MRS in clinical biology has been developed only recently. The in vitro high resolution MRS of body fluids, biopsy samples and surgery specimens has, however, demonstrated exciting potentialities as a multiparametric (full "biochemical profile" analysis), fast, and relatively inexpensive analytical method. Assisted by pattern recognition methods, MR spectra have already provided clinically relevant information in oncology, when the lack of highly specific and sensitive markers has to be deplored for some cancer types.

Proton NMR spectroscopy of plasma lipoproteins: a marker of the immune function in cancer disease?

1H-NMR spectroscopy of cancer plasma statistically detects significant narrowing of the methyl and methylene line widths. This change is due to relative increase in light density lipoproteins (VLDL and LDL) compared to heavy density lipoproteins (HDL). This observation had raised great hopes for a simple and universal screening test of cancer patients. Furthermore, the same signal can be observed in the plasma of pregnant women and heart transplanted patients undergoing an immunosuppressive treatment. This signal disappears after child's birth and during graft rejection processes. These observations suggest that the test initially proposed by Fossel in 1986 reveals a specific immunological status developed by the organism in "symbiosis" with "foreign" cells, rather than a cancerous disease.

Magnetic resonance spectroscopy in cancer: phospholipid, neutral lipid and lipoprotein metabolism and function.

A Special Symposium on "Lipid Metabolism and Function in Cancer. Significance of Magnetic Resonance Spectroscopy (MRS) Measurements in Relation to Biochemical Processes and Cellular Control" was held by the EU BIOMED 1 Concerted Action for "Cancer and Brain Disease Characterization and Therapy Assessment by Quantitative MRS", within the 5th International Conference of Anticancer Research (Corfu, 18-20 October 1995). The aims of the Symposium were to discuss present knowledge and problems in the field of phospholipid, neutral lipid and lipoprotein metabolism in cancer, together with the new potential of these inter-linked areas of research arising from advanced in vitro and in vivo MRS methodologies. This guest editorial is intended to introduce the reader of the Proceedings published in this issue, into the stimulating atmosphere of the Symposium, by summarizing some of the exciting proposals, views, ideas and open questions which emerged from presentations and multi-disciplinary, plenary discussions.

Proton NMR spectroscopy of plasma lipoproteins in the experimental Lewis lung carcinoma.

Plasma and fractionated lipoproteins from 40 Lewis Lung Carcinoma grafted mice were tested from the first day up to the fatal issue by biochemical analyses and water suppressed 1H NMR spectroscopy. We have confirmed first, that the 1H NMR spectra of plasma lipoproteins are modified by the tumoral state and could provide a useful marker of the disease as long as they are used for individual follow-up with appropriate spectral parameters. Using fractionated lipoproteins we have demonstrated secondly, that the observed spectral modifications do not result from a specific cancer lipoprotein but from quantitatively modified ratio between Very Light Density Lipoproteins and High Density Lipoproteins.

Proton NMR studies of tissue water phases during chemical carcinogenesis in rats.

The water content, relative ratio of bound water, mean 1H NMR spin-lattice relaxation time T1, and T1 of bound water fraction were studied on rat liver during the course of cancer induction by diethylnitrosamine. Using the FETS model proposed by Fung, the results were discussed according to histology. Liver T1 increase was correlated with water content and a regular decrease of T1b was observed during the course of hepatocarcinogenesis, associated with a shift of the position of the minimum of T1b toward the negative temperatures. A biphasic decay of T1b was also noticed in the presence of hepatocarcinoma nodules.

Proton nuclear magnetic resonance spectroscopy of plasma lipoprotein: technical problems and potential interest in cancer disease.

This paper reviews several methods presently available for analysing lipoprotein NMR spectra. Two main steps can be distinguished: NMR signal processing and data analysis. Time domain (wavelet transform) and frequency domain (curve fitting) signal processing methods are compared. Statistical methods of data analysis (Ascending Hierarchical Classification, Correspondence Analysis and Principal Component Analysis) have been tested on simulated NMR data of plasma lipoprotein with different numbers of sampling points and different noise levels. These few examples clearly attest that the NMR approach to complex "mixture" (such as body fluids) analysis is emerging from its infancy. New interest in plasma lipoprotein analysis in cancer biology is finally discussed in the light of previous clinical and experimental results and of understanding of lipid metabolism in cancer.

Detection of early gamma-postirradiation effects in murine spleen by proton NMR relaxation times.

BACKGROUND: It was our aim to evaluate the potential of proton relaxation times for the early detection of radiation-induced spleen changes. MATERIALS AND METHODS: Female Swiss mice were irradiated with doses ranging from 0.05 Gy to 4 Gy. The body weight, the spleen weight and the spleen water content of single animals were determined. Measurements of longitudinal (T1) and transversal (T2) proton relaxation times of the spleen samples were performed in a 0.47 T spectrometer. Histological examinations of the control and irradiated organs were performed. RESULTS: NMR measurements during the first five days after irradiation showed that total body gamma-irradiation with doses from 1.5 Gy to 4 Gy results in decreasing T1 of the murine spleen. Significant shortening in T2 was observed for the spleen of animals irradiated with a dose of 4 Gy. Histological examinations demonstrated subnormal architecture in slices derived from animals irradiated with 2 Gy and 4 Gy. CONCLUSION: The fluctuations of the spleen T1 and T2 of irradiated mice are correlated with relative spleen weight and can be used to estimate radiation induced changes in this organ.