Poonam Rana

Nuclear magnetic resonance spectroscopy-based metabonomic investigation of biochemical effects in serum of γ-irradiated mice

Purpose: Radiation exposure induces change in many biological compounds. It is important to assess the physiological and biochemical response to an absorbed dose of ionising radiation due to intentional or accidental event and to predict medical consequences for medical management. In the present study, nuclear magnetic resonance (NMR) spectroscopy-based metabolic profiling was used in mice serum for identification of radiation-induced changes at metabolite level. Materials and methods: Mice were irradiated with 3, 5 and 8 Gray of γ-radiation dose and serum samples collected at day 1, 3 and 5 post irradiation were analysed by proton nuclear magnetic resonance (1H NMR) spectroscopy. 1H NMR spectra of serum were analysed by pattern recognition using principal component analysis. Results: Irradiated mice serum showed distinct metabonomic phenotypes and revealed dose- and time-dependent clustering of irradiated groups. 1H NMR spectral analysis exhibited increased lactate, amino acids, choline and lipid signals as well as decreased glucose signals. These findings indicate radiation-induced disturbed energy, lipid and protein metabolism. Conclusions: The information obtained from this study reflects multiple physiological dysfunctions. The study promises the application of NMR-based metabonomics in the field of radiobiology, for development of metabolic-based markers for screening of risk populations and medical management in these cases.

Radiation‐induced early changes in the brain and behavior: Serial diffusion tensor imaging and behavioral evaluation after graded doses of radiation

The nuclear arsenal and the use of nuclear technologies have enhanced the likelihood of whole‐body/partial‐body radiation exposure. The central nervous system is highly susceptible to even low doses of radiation. With the aim of detecting and monitoring the pathologic changes of radiation‐induced damage in brain parenchyma, we used serial diffusion tensor magnetic resonance imaging (DTI) with a 7T magnetic resonance unit and neurobehavioral assessments mice irradiated with 3‐, 5‐, and 8‐Gy doses of radiation. Fractional anisotropy (FA) and mean diffusivity (MD) values at each time point (baseline, day 1, day 5, and day 10) were quantified from hippocampus, thalamus, hypothalamus, cudate‐putamen, frontal cortex, sensorimotor cortex, corpus callosum, cingulum, and cerebral peduncle. Behavioral tests were performed at baseline, day 5, and day 10. A decrease in FA values with time was observed in all three groups. At day 10, dose‐dependent decreases in FA and MD values were observed in all of the regions compared with baseline. Behavioral data obtained in this study correlate with FA values. Radiation‐induced affective disorders were not radiation dose dependent, insofar as the anxiety‐like symptoms at the lower dose (3 Gy) mimics to the symptoms with the higher dose (8 Gy) level but not with the moderate dose. However, there was a dose‐dependent decline in cognitive function as well as FA values. Behavioral data support the DTI indices, so it is suggested that DTI may be a useful tool for noninvasive monitoring of radiation‐induced brain injury.

Neurodegenerative evidences during early onset of depression in CMS rats as detected by proton magnetic resonance spectroscopy at 7 T

Depression is a complex psychiatric disorder characterized by anhedonia and feeling of sadness and chronic mild stress (CMS) seems to be a valuable animal model of depression. CMS animal model was induced and validated using behavioral studies. In the present study we investigated the neuro-metabolite changes occurring in prefrontal cortex and hippocampus during the onset of depression, in CMS rat model using in vivo proton magnetic resonance spectroscopy ((1)H MRS) at field strength of 7 T. Results showed that CMS caused depression-like behavior in rats, as indicated by the decrease in sucrose consumption and locomotor activity. (1)H MRS was performed in both control and CMS rats (n=10, in each group) and the quantitative assessment of the neurometabolites was done using LC model. Relative concentrations of all the metabolites along with the macromolecules were calculated for analysis. The results revealed a significant decrease of glutamate (Glu), glutamine (Gln), NAA+NAAG, Glx and GABA levels in both hippocampus and prefrontal cortex of CMS animals and an elevated level of myo-ionisitol (mI) and taurine (Tau) was observed only in hippocampus. These metabolite fluctuations revealed by proton MRS indicate that there might be change in the neuronal integrity of the glial cells and neurons within prefrontal cortex and hippocampus in CMS model of depression. The present study also suggests that there may be a degenerative process concerning the brain morphology in the CMS rats. The overall finding using (1)H MRS suggests that, there might be a major role of the glia and neuron in the onset of depression.

Bio-energetic impairment in human calf muscle in thyroid disorders: a 31P MRS study

Mitochondrial metabolism particularly oxidative phosphorylation is greatly influenced by thyroid hormones. Earlier studies have described neuromuscular symptoms as well as impaired muscle metabolism in hypothyroid and hyperthyroid patients. In this study, we intend to look in to the muscle bioenergetics including phosphocreatine recovery kinetics based oxidative metabolism in thyroid dysfunction using in vivo (31)P nuclear magnetic resonance spectroscopy (MRS). (31)P MRS was carried out at resting state on 32 hypothyroid, 10 hyperthyroid patients and 25 control subjects. Nine out of 32 hypothyroid patients and 17 out of 25 control subjects under went exercise protocol for oxidative metabolism study and performed plantar flexion exercise while lying supine in 1.5 T magnetic resonance scanner using custom built exercise device. MRS measurements of inorganic phosphate (Pi), phosphocreatine (PCr), phosphodiesters (PDE) and adenosine triphosphate (ATP) of the calf muscle were acquired during rest, exercise and recovery phase. PCr recovery rate constant (k(PCr)) and oxidative capacity were calculated by monoexponential fit of PCr versus time (t) at the beginning of recovery. During resting condition in hypothyroid patients, PCr/Pi ratio was reduced whereas PDE/ATP and Pi/ATP were increased. However, in case of hyperthyroidism, an increased PCr/Pi ratio and reduced PDE/ATP and Pi/ATP were observed. The results confirmed differential energy status of the muscle due to increased or decreased levels of thyroid hormone. Our results also demonstrate reduced oxidative metabolism in hypothyroid patients based on PCr recovery kinetics. PCr recovery kinetics study after exercise revealed decreased PCr recovery rate constant (k(PCr)) in hypothyroid patients compared to controls that resulted in decrease in oxidative capacity of muscle by 50% in hypothyroids. These findings are consistent with a defect of high energy phosphate mitochondrial metabolism in thyroid dysfunction.

Assessment of the metabolic profile in Type 2 diabetes mellitus and hypothyroidism through proton MR spectroscopy

The metabolic changes in the brain of patients affected with Type 2 diabetes mellitus (DM) alone, both Type 2 DM and hypothyroidism and hypothyroidism only were investigated using proton magnetic resonance spectroscopy ((1)H MRS). Single-voxel spectroscopy was carried out in right and left frontal lobe white matter, left parietal white matter and left occipital gray matter. Choline (Cho)/creatine (Cr) value was found to be increased in the left occipital gray matter of the subjects affected with Type 2 DM and both Type 2 DM and hypothyroidism as compared to controls. No significant change in the Cho/Cr value in the occipital gray matter was observed in hypothyroid subjects as compared to controls. However, they showed an increased level of Cho/Cr in the frontal white matter. High Cho is associated with altered membrane phospholipid metabolism. The high Cho in frontal white matter in hypothyroids and occipital gray matter in diabetic patients suggests that, though both the diseases are endocrine disorders, they differ from each other in terms of regional brain metabolite changes.

Demyelinating evidences in CMS rat model of depression: A DTI study at 7 T

Depression is among the most debilitating diseases worldwide. Long-term exposure to stressors plays a major role in development of human depression. Chronic mild stress (CMS) seems to be a valid animal model for depression. Diffusion tensor imaging (DTI) is capable of inferring microstructural abnormalities of the white matter and has shown to serve as non-invasive marker of specific pathology. We developed a CMS rat model of depression and validated with behavioral experiments. We measured the diffusion indices (mean diffusivity (MD), fractional anisotropy (FA), axial (λ∥) and radial (λ⊥) diffusivity) to investigate the changes in CMS rat brain during depression onset. Diffusion indices have shown to be useful to discriminate myelin damage from axon loss. DTI was performed in both control and CMS rats (n=10, in each group) and maps of FA, MD, λ∥ and λ⊥ diffusivity values were generated using in-house built software. The diffusion indices were calculated by region of interest (ROI) analysis in different brain regions like the frontal cortex, hippocampus, hypothalamus, cingulum, thalamus, caudate putamen, corpus callosum, cerebral peduncle and sensory motor cortex. The results showed signs of demyelination, reflected by increased MD, decreased FA and increased λ⊥. The results also suggest a possible role of edema or inflammation concerning the brain morphology in CMS rats. The overall finding using DTI suggests there might be a major role of loss of myelin sheath, which leads to disrupted connectivity between the limbic area and the prefrontal cortex during the onset of depression. Our findings indicate that interpretation of these indices may provide crucial information about the type and severity of mood disorders.

Glutamate level in anterior cingulate predicts anxiety in healthy humans: A magnetic resonance spectroscopy study

Anxiety, a personality dimension in healthy humans, has been found to be associated with many functional consequences such as increased distractibility and attentional bias in favour of threat-related information, along with morphological and microstructural changes in the brain. The associated metabolic/neurochemical alterations are sparsely studied. In the present magnetic resonance spectroscopy (MRS) study, we investigated the possible relationship between regional brain chemistry within anterior cingulate cortex (4-cm(3) voxel) and hippocampus (2.5-cm(3) voxel) and anxiety (measured by State-Trait Anxiety Inventory) in our subject group. In the anterior cingulate cortex, multivariate analysis of covariance showed an increase in myo-inositol and combined glutamate and glutamine levels in the high anxiety subject group as compared with the low anxiety group. In the partial correlation analysis between neurochemicals and anxiety, glutamate and combined glutamate and glutamine also showed a predictive value for anxiety. On analysing the trait anxiety sub-score separately, we found glutamate, inositol and combined glutamate and glutamine levels to be increased in the high trait anxiety group as compared with the low trait anxiety group. All three resonances also had a predictive value for trait anxiety. In the hippocampus, none of the neurochemicals showed significant difference between high and low anxiety groups. The study provides a first account of alterations in anterior cingulate cortex neurochemistry in relation to anxiety in healthy subjects. The study thus contributes to the limited literature available on altered metabolism and neural mechanisms underlying sub-clinical anxiety.

Differential biochemical response of rat kidney towards low and high doses of NiCl2 as revealed by NMR spectroscopy

Heavy metals are known for their associated nephrotoxicity and nickel is no exception. An integrated metabonomic approach, based on high‐resolution 1H NMR spectroscopy, was applied to determine the acute biochemical effects of NiCl2 on the renal tissues of rats. Kidney homogenates from rats treated with NiCl2 at two dose levels (4 and 20 mg kg−1 b.w., i.p.) and those from controls were analysed using 1H NMR spectroscopy and also assessed for antioxidant parameters at days 1, 3 and 5 post‐dose. The major metabolite changes corresponding to nickel exposure were related to amino acids, osmolytes and energy metabolites. Differential responses were observed in 1H NMR spectra with exposure to low and high doses of NiCl2. For high doses, 1H NMR spectral analysis revealed alterations in renal tissues, along with damage to the cortical and papillary region and depletion of renal osmolytes such as betaine, trimethyl amine oxide, myo‐inositol and taurine, which persisted until day 5 post‐dose. The metabolite profile of 1H NMR spectra obtained from animals treated with lower dose of NiCl2 initially increased as an immediate stress response and then showed signs of recovery with the passage of time. NMR spectral analysis was well corroborated with histopathological and oxidative stress results. Nickel‐induced oxidative stress was observed in both groups of animals with increased levels of antioxidant parameters at initial time points, but continued to increase in the high‐dose group. The present study shows a huge potential of metabonomics for mapping organ‐based metabolic response during heavy metal toxicity. Copyright

Effect of vitamin D supplementation on muscle energy phospho-metabolites: a 31P magnetic resonance spectroscopy-based pilot study

Abstract There are several published reports on the prevalence of low vitamin D levels in otherwise healthy Indian population. Vitamin D deficiency has shown variable effect on muscle performance and strength but there is paucity of data on the effect of vitamin D deficiency on muscle energy metabolism. The present study was proposed to investigate the influence of severe vitamin D deficiency on high-energy metabolite levels in resting skeletal muscle and thereafter, monitor the response after vitamin D supplementation using 31P magnetic resonance spectroscopy (MRS). Study was conducted on 19 otherwise healthy subjects but with low serum 25(OH)D levels (<5 ng/ml). Subjects were supplemented with cholecalciferol at a dose of 60 000 IU/week for 12 weeks. MRS measurements of inorganic phosphate (Pi), phosphocreatine (PCr), phosphodiester (PDE) and ATP of the calf muscle were taken pre- and post-vitamin D supplementation. The study revealed significantly increased PCr/Pi ratio and decreased [Pi] and PDE/ATP ratio with raised serum 25(OH)D levels after 12 weeks of supplementation. The study indicates that serum 25(OH)D level plays an important role in improving the skeletal muscle energy metabolism and vitamin D deficiency might be one of the primary reasons for prevalence of low PCr/Pi ratio and high PDE values in normal Indian population as reported earlier. The findings of this preliminary study are highly encouraging and warrant further in-depth research, involving larger number of subjects of different age groups, regions and socio-economic sections of the society to further strengthen a correlation between vitamin D levels and muscle energy metabolism.

Study of acute biochemical effects of thallium toxicity in mouse urine by NMR spectroscopy.

Thallium (Tl) is a toxic heavy metal and its exposure to the human body causes physiological and biochemical changes due to its interference with potassium‐dependent biological reactions. A high‐resolution 1H NMR spectroscopy based metabonomic approach has been applied for investigating acute biochemical effects caused by thallium sulfate (Tl2SO4). Male strain A mice were divided in three groups and received three doses of Tl2SO4 (5, 10 and 20 mg kg−1 b.w., i.p.). Urine samples collected at 3, 24, 72 and 96 h post‐dose time points were analyzed by 1H NMR spectroscopy. NMR spectral data were processed and analyzed using principal components analysis to represent biochemical variations induced by Tl2SO4. Results showed Tl‐exposed mice urine to have distinct metabonomic phenotypes and revealed dose‐ and time‐dependent clustering of treated groups. The metabolic signature of urine analysis from Tl2SO4‐treated animals exhibited an increase in the levels of creatinine, taurine, hippurate and β‐hydroxybutyrate along with a decrease in energy metabolites trimethylamine and choline. These findings revealed Tl‐induced disturbed gut flora, membrane metabolite, energy and protein metabolism, representing physiological dysfunction of vital organs. The present study indicates the great potential of NMR‐based metabonomics in mapping metabolic response for toxicology, which could ultimately lead to identification of potential markers for Tl toxicity. Copyright