Reduction In Adenosine Triphosphate Levels Research Articles

Urolithin a Improves Motor Dysfunction Induced by Copper Exposure in SOD1G93A Transgenic Mice Via Activation of Mitophagy.

Amyotrophic lateral sclerosis (ALS) is a neurodegenerative disease pathologically characterized by selective degeneration of motor neurons resulting in a catastrophic loss of motor function. The present study aimed to investigate the effect of copper (Cu) exposure on progression of ALS and explore the therapeutic effect and mechanism of Urolithin A (UA) on ALS. 0.13 PPM copper chloride drinking water was administrated in SOD1G93A transgenic mice at 6weeks, UA at a dosage of 50mg/kg/day was given for 6weeks after a 7-week Cu exposure. Motor ability was assessed before terminal anesthesia. Muscle atrophy and fibrosis, motor neurons, astrocytes and microglia in the spinal cord were evaluated by H&E, Masson, Sirius Red, Nissl and Immunohistochemistry Staining. Proteomics analysis, Western blotting and ELISA were conducted to detect protein expression. Mitochondrial adenosine triphosphate (ATP) and malondialdehyde (MDA) levels were measured using an assay kit. Cu-exposure worsened motor function, promoted muscle fibrosis, loss of motor neurons, and astrocyte and microglial activation. It also induced abnormal changes in mitochondria-related biological processes, leading to a significant reduction in ATP levels and an increase in MDA levels. Upregulation of P62 and downregulation of Parkin, PINK1, and LAMP1 were revealed in SOD1G93A mice with Cu exposure. Administration of UA activated mitophagy, modulated mitochondria dysfunction, reduced neuroinflammation, and improved gastrocnemius muscle atrophy and motor dysfunction in SOD1G93A mice with Cu exposure. Mitophagy plays critical role in ALS exacerbated by Cu exposure. UA administration may be a promising treatment strategy for ALS.

Characterization of reduced astrocyte creatine kinase levels in Alzheimer's disease.

The creatine-phosphocreatine cycle serves as a crucial temporary energy buffering system in the brain, regulated by brain creatine kinase (CKB), in maintaining Adenosine triphosphate (ATP)levels. Alzheimer's disease (AD) has been linked to increased CKB oxidation and loss of its regulatory function, although specific pathological processes and affected cell types remain unclear. In our study, cerebral cortex samples from individuals with AD, dementia with Lewy bodies (DLB), and age-matched controls were analyzed using antibody-based methods to quantify CKB levels and assess alterations associated with disease processes. Two independently validated antibodies exclusively labeled astrocytes in the human cerebral cortex. Combining immunofluorescence (IF) and mass spectrometry (MS), we explored CKB availability in AD and DLB cases. IF and Western blot analysis demonstrated a loss of CKB immunoreactivity correlated with increased plaque load, severity of tau pathology, and Lewy body pathology. However, transcriptomics data and targeted MS demonstrated unaltered total CKB levels, suggesting posttranslational modifications (PTMs) affecting antibody binding. This aligns with altered efficiency at proteolytic cleavage sites indicated in the targeted MS experiment. These findings highlight that the proper function of astrocytes, understudied in the brain compared with neurons, is highly affected by PTMs. Reduction in ATP levels within astrocytes can disrupt ATP-dependent processes, such as the glutamate-glutamine cycle. As CKB and the creatine-phosphocreatine cycle are important in securing constant ATP availability, PTMs in CKB, and astrocyte dysfunction may disturb homeostasis, driving excitotoxicity in the AD brain. CKB and its activity could be promising biomarkers for monitoring early-stage energy deficits in AD.

Mitochondrial dysfunction as a target in spinal cord injury: intimate correlation between pathological processes and therapeutic approaches.

Traumatic spinal cord injuries interrupt the connection of all axonal projections with their neuronal targets below and above the lesion site. This interruption results in either temporary or permanent alterations in the locomotor, sensory, and autonomic functions. Damage in the spinal tissue prevents the re-growth of severed axons across the lesion and their reconnection with neuronal targets. Therefore, the absence of spontaneous repair leads to sustained impairment in voluntary control of movement below the injury. For decades, axonal regeneration and reconnection have been considered the opitome of spinal cord injury repair with the goal being the repair of the damaged long motor and sensory tracts in a complex process that involves: (1) resealing injured axons; (2) reconstructing the cytoskeletal structure inside axons; (3) re-establishing healthy growth cones; and (4) assembling axonal cargos. These biological processes require an efficient production of adenosine triphosphate, which is affected by mitochondrial dysfunction after spinal cord injury. From a pathological standpoint, during the secondary stage of spinal cord injury, mitochondrial homeostasis is disrupted, mainly in the distal segments of severed axons. This result in a reduction of adenosine triphosphate levels and subsequent inactivation of adenosine triphosphate-dependent ion pumps required for the regulation of ion concentrations and reuptake of neurotransmitters, such as glutamate. The consequences are calcium overload, reactive oxygen species formation, and excitotoxicity. These events are intimately related to the activation of necrotic and apoptotic cell death programs, and further exacerbate the secondary stage of the injury, being a hallmark of spinal cord injury. This is why restoring mitochondrial function during the early stage of secondary injury could represent a potentially effective therapeutic intervention to overcome the motor and sensory failure produced by spinal cord injury. This review discusses the most recent evidence linking mitochondrial dysfunction with axonal regeneration failure in the context of spinal cord injury. It also covers the future of mitochondria-targeted therapeutical approaches, such as antioxidant molecules, removing mitochondrial anchor proteins, and increasing energetic metabolism through creatine treatment. These approaches are intended to enhance functional recovery by promoting axonal regeneration-reconnection after spinal cord injury.

Mitochondrial Diabetes is Associated with tRNALeu(UUR) A3243G and ND6 T14502C Mutations.

BackgroundMutations in mitochondrial DNA (mtDNA) are associated with type 2 diabetes mellitus (T2DM). In particular, m.A3243G is the most common T2DM-related mtDNA mutation in many families worldwide. However, the clinical features and pathophysiology of m.A3243G-induced T2DM are largely undefined.MethodsTwo pedigrees with maternally inherited T2DM were underwent clinical, molecular and biochemical assessments. The mtDNA genes were PCR amplified and sequenced. Mitochondrial adenosine triphosphate (ATP) and reactive oxygen species (ROS) were measured in polymononuclear leukocytes derived from three patients with both the m.A3243G and m.T14502C mutations, three patients with only the m.A3243G mutation and three controls without these mutations. Moreover, GJB2, GJB3 and GJB6 mutations were screened by PCR-Sanger sequencing.ResultsMembers of the two pedigrees manifestated variable clinical phenotypes including diabetes and hearing and vision impairments. The age at onset of T2DM varied from 31 to 66 years, with an average of 41 years. Mutational analysis of mitochondrial genomes indicated the presence of the m.A3243G mutation in both pedigrees. Matrilineal relatives in one of the pedigrees harbored the coexisting of m.A3243G and m.T14502C mutations. Remarkably, the m.T14502C mutation, which causes the substitution of a conserved isoleucine for valine at position 58 in ND6 mRNA, may affect the mitochondrial respiratory chain functions. Biochemical analysis revealed that cell lines bearing both the m.A3243G and m.T14502C mutations exhibited greater reductions in ATP levels and increased ROS production compared with those carrying only the m.A3243G mutation. However, we did not find any mutations in the GJB2, GJB3 and GJB6 genes.ConclusionOur study indicated that mitochondrial diabetes is associated with the tRNALeu(UUR) A3243G and ND6 T14502C mutations.

The Clinical Benefit of D-ribose in Cardiovascular Ischemic Disease: A New Potential Energy Supplement

Cardiovascular disease is still the leading cause of death worldwide. Atherosclerosis is the most common type of cardiovascular disease and can be caused by various factors, such as genetics and lifestyle. Cells need energy to maintain their integrity and function. Energy that is often used is adenosine triphosphate (ATP). Atherosclerosis can cause myocardial ischemia and induced reduction of ATP levels, so that that not only affects cellular energy, but also alters the normal function. D-ribose is a pentose carbohydrate that has been shown to increase cellular energy levels and improve function after ischemia in pre-clinical studies. It has shown potential benefits in clinical evaluation. This article aims to explain the role of D-ribose in increasing energy levels and myocardial function in ischemic cardiovascular disease.

Aldosterone stimulation mediates cardiac metabolism remodeling via Sirt1/AMPK signaling in canine model.

Aldosterone (Aldo), a pivotal hormone that is ubiquitously expressed in systemic tissues of mammals, is a crucial factor in the pathogenesis of cardiac disease. Accumulating evidence suggests that disturbances in cell energy metabolism are involved in increasing aldosterone levels. However, the precise mechanism underlying the impact of cardiac metabolic remodeling underlying aldosterone stimulation remains limited. In this work, we evaluated the underlying effect of aldosterone on regulating cardiac metabolism remodeling in a canine model. Fifteen beagle dogs were divided into a control group (n = 5), Aldo group (n = 5), and a group treated with spironolactone (SP), a mineralocorticoid receptor antagonist (n = 5), for 4weeks. Blood pressure, electrocardiogram and respiratory parameters, H&E, Masson staining, ultrastructural changes, the adenosine triphosphate (ATP) and free fatty acid (FFA) levels of ventricular tissues, the level of mRNA, and the protein expression of key metabolic factors and regulators were assessed. The Sirt1/AMPK signaling pathway was significantly inhibited in the canine model of aldosterone stimulation, resulting in a reduction of the key downstream metabolic factors involved in glucose and fatty acid oxidation. The dysregulation of expression of key factors in glycogen metabolism led to glycogen deposition, an increase in FFA levels, a reduction in ATP levels, apoptosis, inflammatory cell infiltration, and mitochondrial damage in the ventricular myocardium. These effects were significantly restored by spironolactone. Aldosterone stimulation induced cardiac metabolic remodeling in ventricular cardiomyocytes possibly through the Sirt1/AMPK signaling pathway, implying that this pathway may provide a novel therapeutic target for cardiac metabolic remodeling.

The antidepressant clomipramine induces programmed cell death in Leishmania amazonensis through a mitochondrial pathway.

Despite many efforts, the currently available treatments for leishmaniasis are not fully effective. To discover new medications, drug repurposing arises as a promising strategy. We present data that supports the use of the antidepressant clomipramine against Leishmania amazonensis. The drug presented selective activity at micromolar range against both the parasite forms and stimulated nitric oxide production in host macrophages. Regarding the mechanism of action, clomipramine led parasites do mitochondrial depolarization, which coupled with the inhibition of trypanothione reductase induced strong oxidative stress in the parasites. The effects observed in promastigotes included lipoperoxidation, plasma membrane permeabilization, and apoptosis hallmarks (i.e., DNA fragmentation, phosphatidylserine exposure, and cell shrinkage). The mechanism of action in both parasitic forms was quite similar, but amastigotes also exhibited energetic stress, reflected by a reduction of adenosine triphosphate levels. Such differential effects might be attributable to the metabolic particularities of each form of the parasitic. Ultrastructural alterations of the endomembrane system and autophagy were also observed, possibly indicating an adaptive response to oxidative stress. Our results suggest that clomipramine interferes with the redox metabolism of L. amazonensis. In spite of the cellular responses to recover the cellular homeostasis, parasites underwent programmed cell death.

Research on energy metabolism, lignification and veil opening in postharvest white mushroom ( Agaricus bisporus ) under high O 2 /CO 2 controlled atmospheres (HOC-CA)

White mushroom is an abundant nutritional but perishable product; veil opening and stipe lignification are two of the obvious characte-ristics of senescence. In order to study the relationship between the energy charge, veil opening and lignification of the white mushroom, high O2/CO2 controlled atmospheres (HOC-CA) were utilized at 2±1 °C. The treatments comprised the following 100% O2, 80% O2 + 20% CO2, 60% O2 + 40% CO2, 40% O2 + 60% CO2 respectively, with air as the control. In this study, sensorial and physiological qualities, adenosine triphosphate (ATP) content, energy charge level, lignin content, phenylalanine ammonia-lyase (PAL), cinnamyl alcohol dehydrogenase (CAD) and peroxidase (POD) activity were studied. The optimal condition 80% O2 + 20% CO2 treatment inhibited both respiration rate and veil opening of the mushroom; suppressed PAL, POD and CAD activity. It also retarded the reduction of ATP level, energy charge level, and growth of lignin content. Exogenous ATP had an effect on inhibiting veil opening and lignification. These results indicated that there was a negative correlation between the ATP content, cap opening and stipe lignification. Appropriate HOC-CA maintained high energy, preserved the white mushroom well and extended the storage time to 24 days.

Implementation of Cleaning Process for Mobile Patient Equipment

Background Mobile patient equipment (MPE) such as Dynamap machines (i.e., blood pressure monitoring devices, thermometer and pulse-oximeter), ultrasound machines, electrocardiogram (EKG) bladder scanners and language line translator phones may be significant fomites for the transfer of infections between patients in acute care settings. Baseline adenosine triphosphate (ATP) data from a Level 1 Trauma Center suggested that MPE are not cleaned regularly between patients. The objective of this quality improvement project was to implement a standardized, effective cleaning process for MPE and monitor success of implementation through the use of ATP monitoring and real-time data feedback.Methods A detailed cleaning process and schedule was developed MPE. Education was provided to staff on between use cleaning and an extensive cleaning process to be performed once daily. Cleanliness of MPE was tested through weekly ATP monitoring and the results were provided to floor educators and managers. Median ATP and passing rate were assessed. ATP pass/fail cut-off was set according to manufacturer’s recommendations. Passing ATP level was <250 relative light units (RLU), intermediate level was 250–500 RLU and failing level was >500 RLU.ResultsThe overall median ATP level of all MPE decreased from 755 RLU (N = 102) to 236 RLU (N = 425) 16 weeks post-intervention (Figure 1). The pass rate increased from 19.6% to 52.0%. The blood pressure cuff ATP level demonstrated a 78% decrease from 969.5 RLU (N = 12) to 219 RLU (N = 84). The pulse-ox ATP level also decreased by 78% from 1884 RLU (N = 9) to 407 RLU (N = 86) post-intervention. An 84% reduction in ATP level was identified in the language line translator phone (1,284 RLU to 198 RLU).Conclusion Sixteen weeks post-implementation of this quality improvement project demonstrated that patient equipment is consistently being cleaned and the ATP levels are being maintained at a low level. Future directions include broadening the type of equipment that are assessed through ATP, expanding this project to outpatient settings and exploring the sustainability in the absence of ATP data feedback. The improvement of the cleanliness of the equipment potentially has the impact of decreasing infections throughout the hospital.Disclosures All authors: No reported disclosures.

PGC-1α regulates the cell cycle through ATP and ROS in CH1 cells

Peroxisome proliferator-activated receptor-γ coactivator 1α (PGC-1α) is a transcriptional co-activator involved in mitochondrial biogenesis, respiratory capacity, and oxidative phosphorylation (OXPHOS). PGC-1α plays an important role in cellular metabolism and is associated with tumorigenesis, suggesting an involvement in cell cycle progression. However, the underlying mechanisms mediating its involvement in these processes remain unclear. To elucidate the signaling pathways involved in PGC-1α function, we established a cell line, CH1 PGC-1α, which stably overexpresses PGC-1α. Using this cell line, we found that over-expression of PGC-1α stimulated extra adenosine triphosphate (ATP) and reduced reactive oxygen species (ROS) production. These effects were accompanied by up-regulation of the cell cycle checkpoint regulators CyclinD1 and CyclinB1. We hypothesized that ATP and ROS function as cellular signals to regulate cyclins and control cell cycle progression. Indeed, we found that reduction of ATP levels down-regulated CyclinD1 but not CyclinB1, whereas elevation of ROS levels down-regulated CyclinB1 but not CyclinD1. Furthermore, both low ATP levels and elevated ROS levels inhibited cell growth, but PGC-1α was maintained at a constant level. Together, these results demonstrate that PGC-1α regulates cell cycle progression through modulation of CyclinD1 and CyclinB1 by ATP and ROS. These findings suggest that PGC-1α potentially coordinates energy metabolism together with the cell cycle.

Respiratory pathway metabolism and energy metabolism associated with senescence in postharvest Broccoli (Brassica oleracea L. var. italica) florets in response to O2/CO2 controlled atmospheres

Broccoli is a highly perishable vegetable with abundant nutritional and medical value. Recent studies indicated that O2/CO2 controlled atmospheres (CA) was an effective method for broccoli storage.In this study, ATP (adenosine triphosphate) content, energy charge level, respiratory pathway and their key enzymes of broccoli (Brassica oleracea L. var. italica) were investigated, employed CA treatments of 70% O2+30% CO2, 60% O2+40% CO2, 50% O2+50% CO2, 40% O2+60% CO2, 30% O2+70% CO2 at 10°C. The 50% O2+50% CO2 treatment led to an obvious decrease in respiration rate of broccoli and it could inhibit the reduction of ATP level, energy charge level, SDH (succinic dehydrogenase), CCO (cytochrome oxidase), and boost the G-6-PDH (glucose-6-phosphate dehydrogenase)+6-PGDH (6-phosphogluconate dehydrogenas) activity. Treatment of 50% O2+50% CO2 suppressed reduction of the TCA (tricarboxyfic-acid-cycle) and CCP (cytochrome oxidase pathway) rate, and increased the HMP (phosphopentose pathway), while had little impact on the EMP (embden-meyerhof-parnas) rate and PGI (phosphohexose isomerase) activity. Compared to 50% O2+50% CO2 treatment, treatment of 30% O2+70% CO2 had an opposite effect on broccoli. Results indicated that O2/CO2 controlled atmospheres could alter the proportion of respiratory pathway, and 50% O2+ 50% CO2 treatment could enhance the endurance capability of broccoli to high CO2 concentration according to the changes of respiratory pathway and delay in its senescence though relatively higher ATP.

Glucose may attenuate isoflurane-induced caspase-3 activation in H4 human neuroglioma cells.

The commonly used inhaled anesthetic isoflurane has been shown to induce caspase-3 activation. However, the underlying mechanism(s) and targeted intervention(s) remain largely to be determined. Isoflurane may induce caspase-3 activation via causing accumulation of reactive oxygen species (ROS), mitochondrial dysfunction, and reduction in adenosine triphosphate (ATP) levels. Therefore, we performed a hypothesis-generation study to determine whether glucose could attenuate isoflurane-induced caspase-3 activation, ROS accumulation, mitochondrial dysfunction, and ATP reduction in cultured cells. H4 human neuroglioma cells (H4 cells) were treated with 2% isoflurane or the control condition plus saline or 50 mM glucose for 6 or 3 hours. Caspase-3 activation, cell viability, levels of ROS and ATP, and mitochondrial membrane potential were determined at the end of the experiments by Western blot analysis and fluorescence assay. We found that the glucose treatment might attenuate isoflurane-induced caspase-3 activation and reduction of cell viability in H4 cells. Moreover, the glucose treatment mitigated the isoflurane-induced increase in ROS levels and reduction in ATP levels in H4 cells. Unexpectedly, we observed that the glucose treatment might not inhibit the isoflurane-induced decrease in mitochondrial membrane potential in H4 cells. Pending further studies, these results suggested that glucose might attenuate isoflurane-induced caspase-3 activation through a mitochondria-independent reduction in ROS levels and enhancement in ATP levels. These findings have established a system and suggest that it is worth performing more research to further investigate whether glucose can attenuate anesthesia neurotoxicity.

Abstract 483: Docosahexaenoic acid inhibits heat shock protein 90 complex and client proteins by reducing intracellular ATP levels in breast and lung cancer

Abstract A major contributor to carcinogenesis is the molecular chaperone heat shock protein 90 (Hsp90). Hsp90 regulates the proper folding and stabilization of several client proteins, including human epidermal growth factor receptor 2 (ErbB2) and hypoxia-inducible factor 1α (HIF-1α). ErbB2 is a member of the epidermal growth factor receptor (EGFR) family of receptor-proteins, and is a clinical target for the treatment of metastatic cancer, while HIF-1α is often stabilized and overexpressed in several cancers. The ability for Hsp90 to become a mature complex and to properly fold client proteins is dependent on adenosine triphosphate (ATP) binding with the co-chaperone, p23. Specific Hsp90 inhibitors, like geldanamycin, target the Hsp90 ATP-binding site to prevent formation of the mature complex. Although supplementation with docosahexaenoic acid (DHA; C22:6 n-3) has not been shown to specifically target the Hsp90 ATP-binding site, the current study demonstrates a similar effect by DHA causing a reduction in available intracellular ATP levels. DHA is the longest, most unsaturated omega-3 polyunsaturated fatty acid (PUFA) existing in biological membranes and has been shown to possess anti-cancer activity in several cancer models. Studies from our laboratory have shown that DHA enrichment can reduce cancer cell metabolism and intracellular ATP levels in both breast and lung cancer models. In this study, a novel mechanism for the ability of DHA to inhibit the Hsp90 chaperone complex is defined. In vitro analyses of BT-474 human breast ductal carcinoma, as well as the A549 human lung adenocarcinoma cell lines were used to determine the impact of DHA-induced decreases of intracellular ATP levels. We found that the reduction in ATP levels by DHA treatment resulted in significant decreases in the association of Hsp90 and p23 in both cell lines. The decreased association of the Hsp90-p23 complex led to decreased levels of ErbB2 and HIF-1α client proteins, suggesting that DHA can modify Hsp90 chaperone function and attenuate client protein levels. Additionally, similar results were found when employing 2-deoxyglucose (2-DG), a glycolytic inhibitor, which confirms that DHA and 2-DG can disrupt Hsp90 molecular chaperone function by decreasing cellular ATP levels. Consistent with these observations, when using a dose of DHA that did not significantly reduce intracellular ATP levels, no change in the Hsp90-p23 chaperone complex or protein levels of ErbB2 and HIF-1α were seen. These results demonstrate a potential use for dietary intervention to improve cancer therapy in tumors that overexpress Hsp90 client proteins. Citation Format: Michael Mouradian, Irvin V. Ma, Erika D. Vicente, Keith D. Kikawa, Amy M. Chattin, Ronald S. Pardini. Docosahexaenoic acid inhibits heat shock protein 90 complex and client proteins by reducing intracellular ATP levels in breast and lung cancer. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 483. doi:10.1158/1538-7445.AM2014-483

A critical study: Assessment of the effect of silica particles from 15 to 500 nm on bacterial viability

The current opinion on the toxicity of nanomaterials converges on a size-dependent phenomenon showing increasing toxicity with decreasing particle sizes. We demonstrate that SiO2 particles have no or only a mild effect on the viability of five bacterial strains, independently from the particle size. A two-hour exposure to 20 mg L−1 of 15, 50 and 500 nm sized SiO2 particles neither alters bacterial adenosine triphosphate (ATP) levels nor reduces the number of colony forming units (CFU). Additionally, we tested the effect of Al2O3-coated LUDOX®-CL (ACS 20) with a primary particle size of 20 nm. In contrast, these particles caused a significant reduction of ATP levels and CFU. Fluorescence microscopy revealed that ACS 20 induced a pronounced agglomeration of the bacteria, which led to underestimated counts in regard of CFU. Bactericide effects as indicated by decreased ATP levels can be explained by bactericide additives that are present in the ACS 20 suspension.

Chi-square analysis of the reduction of ATP levels in L-02 hepatocytes by hexavalent chromium.

This study explored the reduction of adenosine triphosphate (ATP) levels in L-02 hepatocytes by hexavalent chromium (Cr(VI)) using chi-square analysis. Cells were treated with 2, 4, 8, 16, or 32 µM Cr(VI) for 12, 24, or 36 h. Methyl thiazolyl tetrazolium (MTT) experiments and measurements of intracellular ATP levels were performed by spectrophotometry or bioluminescence assays following Cr(VI) treatment. The chi-square test was used to determine the difference between cell survival rate and ATP levels. For the chi-square analysis, the results of the MTT or ATP experiments were transformed into a relative ratio with respect to the control (%). The relative ATP levels increased at 12 h, decreased at 24 h, and increased slightly again at 36 h following 4, 8, 16, 32 µM Cr(VI) treatment, corresponding to a “V-shaped” curve. Furthermore, the results of the chi-square analysis demonstrated a significant difference of the ATP level in the 32-µM Cr(VI) group (P < 0.05). The results suggest that the chi-square test can be applied to analyze the interference effects of Cr(VI) on ATP levels in L-02 hepatocytes. The decreased ATP levels at 24 h indicated disruption of mitochondrial energy metabolism and the slight increase of ATP levels at 36 h indicated partial recovery of mitochondrial function or activated glycolysis in L-02 hepatocytes.

Chrysophanol induces necrosis through the production of ROS and alteration of ATP levels in J5 human liver cancer cells

Anthraquinone compounds have been shown to induce apoptosis in different cancer cell types. Effects of chrysophanol, an anthraquinone compound, on cancer cell death have not been well studied. The goal of this study was to examine if chrysophanol had cytotoxic effects and if such effects involved apoptosis or necrosis in J5 human liver cancer cells. Chrysophanol induced necrosis in J5 cells in a dose- and time-dependent manner. Non-apoptotic cell death was induced by chrysophanol in J5 cells and was characterized by caspase independence, delayed externalization of phosphatidylserine and plasma membrane disruption. Blockage of apoptotic induction by a general caspase inhibitor (z-VAD-fmk) failed to protect cells against chrysophanol-induced cell death. The levels of reactive oxygen species production and loss of mitochondrial membrane potential (DeltaPsi(m)) were also determined to assess the effects of chrysophanol. However, reductions in adenosine triphosphate levels and increases in lactate dehydrogenase activity indicated that chrysophanol stimulated necrotic cell death. In summary, human liver cancer cells treated with chrysophanol exhibited a cellular pattern associated with necrosis and not apoptosis.

Effect of lactate therapy upon cognitive deficits after traumatic brain injury in the rat

In previous studies, it has been shown that intravenous lactate therapy can improve brain neurochemistry, adenosine triphosphate (ATP) generation and outcome after traumatic brain injury (TBI) in rats. In this study, we examined: (1) four L-lactate concentrations to determine the optimal therapeutic dose post TBI in terms of cognitive function; (2) ATP production after TBI for the L-lactate concentration found to be the optimal dose; (3) the possible production of lactic acidosis with the highest L-lactate concentration tested. Thirty minutes following a fluid percussion injury (FPI) over the left cerebral hemisphere, the animals received an intravenous infusion of 10, 28, 100, or 280 mM L-lactate (n = 10 for each group) for 3 h at a rate of 0.65 ml/h. Shams and control injured animals received a saline infusion. At 11-15 days post injury, cognitive deficits were examined using the Morris Water Maze (MWM) test. Three groups of rats were used for ATP analysis: shams, injured + saline infusion, and injury + the optimal lactate dose as determined by the MWM (n = 4/group). Additionally, a group receiving 280 mM L-lactate (n = 5) and one receiving a saline infusion (n = 3) were monitored for arterial blood variables and blood pressures. In the MWM test, only the 100 mM L-lactate-treated injured animals showed a significant reduction in cognitive deficits when compared to saline-treated injured animals (p <or= 0.05). In the ATP study, injured animals without treatment had a 53% reduction in ATP level in the ipsilateral cortex, while animals with 100 mM lactate treatment had a 28% reduction. (p <or= 0.05). No lactic acidosis was induced by the intravenous infusion of 280 mM L-lactate. This study indicates that the intravenous infusion of 100 mM L-lactate provided the optimal concentration of the substrate to ameliorate cognitive impairment, probably via the regeneration of ATP following TBI in rats.

Long-term changes in microbial and biochemical parameters in the Central Indian Basin

Natural kilometer-scale spatial variability and seasonal variability in deep-sea sediments at ∼5000 m depth was studied at two reference sites in the Central Indian Basin. Biochemical parameters such as the labile organic matter (LOM) (consisting of carbohydrates, protein and lipids) and total organic matter (TOM) and biological parameters such as total counts of bacteria and adenosine triphosphate (ATP) were estimated after an interval of 54 months. These were compared with nine locations (12–17 km away) where an artificial sediment disturbance was created by a hydraulic benthic disturber. Besides the above-mentioned parameters, extracellular enzymes, alkaline phosphatase, aminopeptidase and lipase were also monitored at these nine locations. Post-disturbance (PoD1) observations were carried out immediately after the benthic disturbance experiment and compared with the pre-disturbance (PreD) pattern. The restoration process in the test site was studied after a period of 44 months (PoD2). There was a drastic reduction in total counts of bacteria, ATP, carbohydrates and lipids accompanied by a two- to four-fold increase in protein content at the undisturbed reference sites after 54 months. A similar reduction in these parameters was observed in the PoD1 compared to the PreD stage at the test site. An initial increase in TOM content and in the activities of three extracellular enzymes in the PoD1 stage was followed by a reduction in TOM, ATP and organic P content and activities of the extracellular enzymes in the PoD2 phase (i.e. after 44 months). Reduction in ATP levels suggests a negative impact of resedimentation on living biomass in the sediments. Although LOM, protein concentrations and the protein/carbohydrate ratio increased in the PoD2 phase relative to the PoD1 phase, the bacterial numbers were below the concentrations in the PreD stage, indicating slow restoration of benthic conditions. Long-term analyses of the indicator parameters thus showed mixed effects of sediment disturbance. A similar pattern at the undisturbed reference location suggests that natural (seasonal) variability outweighed artificial variability caused by the benthic disturbance in the Central Indian Basin and the former is responsible for bringing about changes in deep-sea benthic ecosystem on long-time scales.

Is Warm Retrograde Blood Cardioplegia Better Than Cold for Myocardial Protection?

Background. This study tests the hypothesis that continuous normothermic retrograde blood cardioplegia is superior to cold intermittent blood cardioplegia in protecting the left and right side of the heart transmurally during an extended cross-clamping period. Methods. Twelve anesthetized, open chest dogs were placed on cardiopulmonary bypass and randomized to receive continuous warm (n = 6) or intermittent cold cardioprotection (n = 6) during a 3-hour aortic cross-clamp period. Transmural left ventricular muscle biopsy specimens were taken before the initiation of cardiopulmonary bypass and 90 and 180 minutes after cross-clamping. Right ventricular (RV) biopsy specimens were taken 180 minutes after aortic cross-clamping. Biopsy specimens were analyzed for adenosine triphosphate, creatine phosphate, and lactate levels and for morphologic changes via electron microscopy. Results. At the end of 180 minutes of cardiopulmonary bypass, the adenosine triphosphate contents of endocardial and epicardial halves of the left ventricular myocardium were only slightly degraded in both cardioplegia groups; a significantly greater reduction in adenosine triphosphate levels occurred in the RV of the warm compared with the cold group ( p < 0.02). The difference in creatine phosphate values in the left ventricle between the cold group (35.2 ± 23.4 nmol/mg cardiac protein) and the warm animals (64.4 ± 24.9 nmol/mg cardiac protein) was not statistically significant, but the RV creatine phosphate stores were significantly better preserved in the warm compared with the cold cardioplegia group ( p < 0.02). Lactate levels increased to a similar extent in both groups, but both values rose significantly over baseline ( p < 0.03). Importantly the electron microscopic score of the left ventricle and RV indicated that cells were reversibly and not irreversibly damaged with both cardioplegic protections. Conclusions. These results suggest the following: (1) Chemical arrest is a major contributor of myocardial preservation during diastolic arrest as used in clinical cardiac surgery. (2) Both methods preserve the ultrastructure of the myocytes transmurally during 3 hours of aortic cross-clamping. (3) Both techniques protect the RV and left ventricle; however, to provide optimal protection of the RV, alternated retrograde and antegrade perfusion might be beneficial over retrograde cardioplegia flow alone, in particular with warm cardioplegia. (Ann Thorac Surg 1997;63:98–104)

A tissue culture ischemic device to study eicosanoid release by pheochromocytoma PC12 cultures

In an attempt to search for neuronal models to investigate the molecular pharmacology of central nervous system ischemia, we have focused on PC12 pheochromocytoma cultures which are now popular in neuroscience research. These chromaffinergic transformed cells, originary from the adrenal medulla, synthesize and release catecholamines and, upon treatment with nerve growth factor (NGF), differentiate to a sympathetic phenotype expressing neurites and excitability. To measure eicosanoid production, undifferentiated or NGF-treated PC12 cultures have been exposed for 1 h to a mixture of N 2/CO 2 (95:5%) resulting in hypoxia (5 ± 1% O 2), followed by 1 h reoxygenation (21% O 2) using a special ischemic device. Hypoxia, up to 2 h, was not followed by significant cytotoxicity or significant production of prostaglandin PGE 2. However, upon reoxygenation, a specific release of PGE 2 (2–3 fold over control) was measured. A similar PGE 2-enhanced release could be induced by ‘chemical hypoxia’ using 2-deoxyglucose and oligomycin to reduce cellular adenosine triphosphate (ATP). Anoxia (0.1–1% O 2, 1 h) achieved by a reduction of culture incubation volume and the reduction in ATP level have been found as critical parameters leading to PC12 cells cytotoxicity. These results emphasize the simplicity and applicability of the tissue culture ischemic device proposed to investigate hypoxia and ischemia at a cellular level.