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Biomedical subjects

R Raghupathi

Publications and source records attributed to R Raghupathi.

65 records · Page 4Linked to original sources
PubMed1995

Changes in gene expression following traumatic brain injury in the rat.

This paper reviews changes in gene expression produced by two rodent models of traumatic brain injury: cortical impact injury and fluid-percussion injury. Cortical impact injury produces transient increases in c-fos mRNA expression, which begin as early as 5 min after injury and subsides by 1 day after injury in the cerebral cortex ipsilateral to injury. In addition, AP-1 transcription factor binding is greatly increased in the injured cerebral cortex at 1, 3, and 5 h post-injury. AP-1 binding remains increased for at least 1 day after injury, while SP-1 transcription factor binding activity does not increase. Additional studies have confirmed increases in c-fos mRNA expression in the hippocampus at 30 min, 1 h, and 3 h after injury. These increases in c-fos mRNA in the hippocampus preceded increased levels of NGF mRNA that were detected at 1 and 3 h but not at 30 min following injury. Following fluid-percussion injury, increases in c-fos mRNA can be detected as early as 2 h following injury in the cortex ipsilateral to the site of injury as well as in the hippocampus. Heat-shock protein (hsp72) mRNA is also increased in the ipsilateral cortex and hippocampus following fluid percussion injury. By 24 h post-injury, both c-fos and hsp72 gene expression return to control levels. Severe but not moderate fluid percussion injury produces increased gene expression for glucose-regulated proteins (grp78, grp94) 12 h following injury. Fluid-percussion injury also produces significant increases in expression of both interleukin-1 beta and tumor necrosis factor-alpha in the injured cortex and ipsilateral hippocampus as early as 1 h post-injury, that remains elevated up to 6 h in the injured cortex and hippocampus.

Animals↗
PubMed1995

Cellular responses to experimental brain injury.

Little is known regarding the molecular (genomic) events associated with the pathophysiology of traumatic brain injury (TBI). This review focusses on the experimental efforts to date elucidating the acute alterations in expression of immediate early genes (IEGs), heat shock proteins (HSPs) and cytokines following experimental brain injury. The immediate early genes, c-fos, c-jun and junB were observed to be bilaterally induced in the cortex and hippocampus as early as 5 min following lateral fluid-percussion (FP) brain injury in the rat. While levels of c-fos and junB mRNA returned to control levels by 2h, c-jun mRNA remained elevated up to 6h post-injury. Increased levels of mRNA for the inducible heat-shock protein (hsp72) were observed up to 12h following injury and were restricted to the cortex ipsilateral to the impact site. Mild induction of the glucose-regulated proteins (grp78 and grp94), which share sequence homology with hsp72, was apparent in the ipsilateral cortex. The cytokines IL-1 beta and TNF alpha were induced at 1h following FP brain injury and remained elevated up to 6h post-injury. These data, while indicative of the complex genomic response to TBI, are also suggestive of the trauma-induced activation of multiple signal transduction pathways.

Animals↗
PubMed1993

Preparation and pharmacological evaluation of enantiomers of certain nonoxygenated aporphines: (+)- and (-)-aporphine and (+)- and (-)-10-methylaporphine.

The subject compounds were prepared as a part of a continuing structure-activity study of the contrasting actions (agonism-antagonism) of (+)- and (-)-11-hydroxy-10-methylaporphine at serotonin (5-HT1A) receptors. None of the targeted nonoxygenated aporphine derivatives demonstrated significant activity in assays for any effects at serotonin 5-HT1A receptors. It is concluded that, in the aporphine series, serotonergic agonist or antagonism requires an alkyl group ortho to a phenolic OH group in the A ring.

Animals↗
PubMed1992

Inhibition of phospholipase A2 by cis-unsaturated fatty acids: evidence for the binding of fatty acid to enzyme.

Calcium-dependent phospholipases A2 are markedly inhibited in vitro by cis-unsaturated fatty acids (CUFAs) and to a much lesser extent by trans-unsaturated or saturated fatty acids. Thus, CUFAs may function as endogenous suppressors of lipolysis. To better understand the mechanism of inhibition, kinetic analysis, fluorescence spectroscopy and gel permeation chromatography were employed to demonstrate that CUFAs interact with a highly purified Ca(2+)-dependent phospholipase A2 from Naja mossambica mossambica venom. Arachidonate inhibited hydrolysis of both [1-14C]oleate-labelled, autoclaved Escherichia coli and [1-14C]linoleate-labelled phosphatidylethanolamine in an apparent competitive manner. When subjected to gel permeation chromatography, [3H]arachidonate, but not [3H]palmitate, comigrated with the enzyme. Arachidonic and other CUFAs increased the fluorescence intensity of the enzyme almost 2-fold in a dose-dependent fashion (50 microM = 180% of control); methyl arachidonate was without effect. Saturated fatty acids had only a modest effect on enzyme fluorescence (50 microM = 122% of control). Concentrations of arachidonate that inhibited in vitro enzymatic activity by almost 80% did not alter binding of phospholipase A2 to the E. coli substrate. Collectively, these data demonstrate that, while CUFAs selectively bind to the enzyme, they do not influence phospholipase A2-substrate interaction. Inhibition of in vitro phospholipase A2 activity by CUFAs may be mediated by the formation of an enzymatically inactive enzyme-substrate-inhibitor complex.

Arachidonic Acid↗
PubMed1992

Binding of phenylalkylamine derivatives at 5-HT1C and 5-HT2 serotonin receptors: evidence for a lack of selectivity.

Certain phenylalkylamine derivatives have been considered to bind selectively at 5-HT2 serotonin receptors. It is now recognized that the most widely used derivatives, i.e., 1-(2,5-dimethoxy-4-X-phenyl)-2-aminopropanes where X = Me (DOM), Br (DOB), and I (DOI) (1-3, respectively) also bind at the more recently identified population of serotonin 5-HT1C receptors. The purpose of the present investigation was to determine whether simple phenylalkylamines bind selectively at one population of receptors over the other. An examination of 34 derivatives reveals (i) similar structure-affinity relationships and (ii) a significant correlation (r = greater than 0.9, n = 25) between 5-HT1C and 5-HT2 affinity. None of the compounds included in the present study displayed more than a 10-fold selectivity for one population of these receptors over the other; the results suggest that these compounds (including the widely used 5-HT2 agonists DOB and DOI) are 5-HT1C/5-HT2 agents.

Animals↗
PubMed1990

Inhibition of human phospholipases A2 by cis-unsaturated fatty acids and oligomers of prostaglandin B1.

Inhibition of human phospholipases A2 by cis-unsaturated fatty acids and their oxidative metabolites and/or polymers was studied using partially purified human phospholipases A2 and [1-14C]oleate labelled, autoclaved E. coli as substrate. As previously reported for other phospholipases A2, oleic and arachidonic acids inhibited human synovial fluid phospholipase A2 with IC50s of 15 and 30 microM respectively. Air oxidation of arachidonic acid or hydroxylation of oleic acid (12-hydroxy-oleate) substantially relieved that inhibition. Similarly, the enzymatically oxidatized metabolite of arachidonate, prostaglandin B1 (PGB1), did not inhibit enzymatic activity. However, prostaglandin Bx (PGBx), an oligomer (n = 6) of PGB1, was a potent inhibitor of Ca(++)-dependent, neutral-active phospholipase A2 activities. Enzymatic activity in acid extracts from human neutrophils, platelets, sperm, plasma, synovial fluid, endometrium, degenerative disc, and snake venom was inhibited by PGBx with IC50s ranging from 0.5-7.0 microM. Inhibition was independent of substrate phospholipid concentration over a 24-fold range (5-120 microM) and PGBx quenched the tryptophan fluorescence of snake venom phospholipase A2 in a dose-dependent manner. Agonist-induced (A23187) release of arachidonic acid from prelabelled human neutrophils and cultured human endothelial cells was also inhibited by PGBx with IC50s of 3 and 20 microM, respectively. These results illustrate that oxidative reactions of cis-unsaturated fatty acids relieve their natural inhibitory activity, and polymerization of an inactive fatty acid metabolite yields a potent inhibitor of in vitro and in situ phospholipase A2 activity.

Arachidonic Acid↗
PubMedDate not supplied

Fatty acid oxidation and myocardial phospholipase A2 activity.

Cis-unsaturated fatty acids, but not saturated fatty acids, inhibited phospholipase A2 activity (PLA2) in vitro, and may function as endogenous suppressors of lipolysis. To probe the possible role of lipid peroxidation in the regulation of myocardial lipid catabolism, a neutral-active and Ca2+-dependent PLA2 was extracted from rat heart and was partially purified by sulfopropyl cation exchange chromatography. Myocardial PLA2 activity was inhibited in a dose-dependent manner by oleic, linoleic, linolenic, and arachidonic acids; the IC50 for arachidonic acid was approx 65 microM. Palmitic acid was not inhibitory. When arachidonic acid was incubated at 37 degrees C, exposed to air, there was a time- and pH-dependent peroxidation of the arachidonic acid as monitored by turbidity, thiobarbituric acid reactivity, and thin layer chromatography. Peroxidation was increased as the pH was lowered from 7.5 to 4.5, and was accompanied by a decrease in PLA2 inhibitory potency. Thus, arachidonate incubated for 24 hours at pH's 4.5, 6.0 and 7.5 lost 84%, 32%, and 20% respectively, of its inhibitory potency. Therefore, in vitro acidosis promotes the oxidation of cis-unsaturated fatty acids and relieves their inhibitory or suppressive activity toward PLA2s. Increased lipid peroxidation of unesterified unsaturated fatty acids during acidosis may therefore promote lipolysis observed during myocardial ischemia and reperfusion injury.

Animals↗
PubMedDate not supplied

TUNEL-positive staining in white and grey matter after fatal head injury in man.

Paraffin sections from the hippocampus, the cingulate gyrus and the insula of 18 head-injured patients who survived between 5 hours and 10 days, and 18 age-matched controls, were stained by the terminal deoxynucleotidyl transferase mediated biotinylated deoxyuridine triphosphate nick end labelling (TUNEL) technique for evidence of in situ DNA fragmentation. Additional staining techniques (HE, combined LFB/CV and immunohistochemistry for GFAP and CD68) were used to characterize any lesions and their time course. Only the occasional TUNEL+ cell per area was seen in the control brains. TUNEL+ cells were identified in both grey and white matter of the head-injured material and their numbers peaked between 24 and 48 hours and were still present at 10 days. Within the hippocampus, fewer TUNEL+ cells were seen in grey (between 3-5 per area) than in the white matter, (up to 51+ per area) whereas in the cingulate gyrus and in the insula, the number of TUNEL+ cells was always greater in the cortex (between 11-20 per area) than in white matter (6-10 per area). In the grey matter, most TUNEL+ cells had the morphology of necrosis. However, the histological appearances of some of the neurons (2-3%), and of oligodendroglia and macrophages in white matter (about 5%) were those of apoptosis.

Adult↗
PubMedDate not supplied

Age does not influence DNA fragmentation in the hippocampus after fatal traumatic brain injury in young and aged humans compared with controls.

Paraffin sections from the hippocampus of 12 head-injured patients (Group A, aged between 4 and 12 years n = 6 and Group B, aged between 64 and 89 years n = 6) and associated age-matched controls were stained by the terminal deoxynucleotidyl transferase-mediated biotinylated deoxyuridine triphosphate nick end labeling (TUNEL) technique for evidence of in-situ DNA fragmentation. TUNEL+ cells were of 2 Types: I (non-apoptotic) and II (apoptotic). In addition sections stained H&E, combined Luxol Fast Blue/Cresyl Violet and by immunohistochemistry for astrocytes (GFAP) and macrophages (CD68) were used to characterize the lesions. Small numbers of Type I TUNEL+ cells were seen in all sectors of the hippocampus except CA2 of both Groups A and B. Type II TUNEL+ cells were mainly found in the white matter. They constituted less than 1% of all TUNEL+ cells. There were similar or fewer TUNEL+ cells in the corresponding areas in the controls compared with the head-injured patients. However, in the dentate fascia and the CA4 sector of the Group B cases, larger numbers of TUNEL+ cells were seen in controls than after trauma. In the grey matter most TUNEL+ cells had the morphology ofnecrosis that corresponded with foci of selective neuronal damage. Only a few TUNEL+ cells were seen in white matter. The occasional Type I TUNEL+ cells were seen in grey matter. It is concluded that the amount and distribution of DNA fragmentation in children and adults is similar and therefore at least in the hippocampus does not provide an explanation for age as an independent variable of outcome after traumatic brain injury in childhood.

Age Factors↗
PubMedDate not supplied

No evidence for the presence of apolipoprotein epsilon4, interleukin-lalpha allele 2 and interleukin-1beta allele 2 cause an increase in programmed cell death following traumatic brain injury in humans.

BACKGROUND: Brain injury after trauma is an important cause of mortality and morbidity in society. There is evidence in both man and laboratory animals that in addition to necrosis, cell loss may occur as a result of programmed cell death (PCD). The cellular and molecular responses after head injury are partly influenced by genetic polymorphisms of apolipoprotein E and the pro-inflammatory cytokine IL-I. AIM: The principal aim of this study was to determine whether the presence of the ApoE epsilon4, IL- 1 alpha2 or IL- 1beta2 allele types influenced the amounts of PCD after head injury compared with controls. METHODS: Paraffin sections from the hippocampus of 38 patients (32 M : 6 F, aged 15 - 75, mean 38 years, survival 7- 576 hours; mean 36 hours) who died after a head injury were stained by Tunel histochemistry and quantified, and genotyping was undertaken by PCR "blind" to clinical detail. RESULTS: There were more Tunel+ cells (neurons and glia) after head injury than in controls with statistically increased numbers in all sectors of the hippocampus including the dentate fascia. However, there was no correlation between ApoEepsilon4, IL- 1 alpha allele 2 and IL- 1beta allele 2 and the amount of Tunel positivity. CONCLUSION: Given that both the ApoE and IL-1 influence outcome after various forms of acute brain injury, further work will be required to determine the mechanism underlying this relationship.

Adolescent↗