Extracellular methemoglobin primes red blood cell aggregation in malaria: An in vitro mechanistic study

Balaji, Shruti; Trivedi, Vishal

doi:10.1016/j.febslet.2012.12.015

Cited by 23 publications

(25 citation statements)

References 45 publications

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“…The level of methemoglobin in malaria patients and in the malaria culture supernatant varies from 0–54 µM [30], [45], [46]. As per the estimate, approximately 200 µM haemozoin is found in P.falciparum infected patients [47] where as the level of haemozin at the brain site may be upto 100 µM [48].…”

Section: Resultsmentioning

confidence: 98%

“…Propogation of malaria parasite within host blood releases a mixture of different pro-oxidant molecules such as methemoglobin, heme, haemozoin (heme polymer) and malaria toxins [5], [6], [7], [30], [47], [48]. Haemozoin is an inert non-biodegradable polymer and macrophages treated with polymer exhibit significant changes in cytokine profile (pro-inflammaory/anti-inflammatory), phagosome maturation, and other functional defects [16], [17], [18], [19], [20], [59].…”

Section: Discussionmentioning

confidence: 99%

“…Malaria culture supernatant was fractionated with ammonium sulfate as described [30]. The different fractions contain variable amount of methemoglobin with fraction P2 contains maximum amount of methemoglobin.…”

Section: Methodsmentioning

confidence: 99%

“…It results in accumulation of parasitized RBCs and pathological complication of brain [26], [27]. Pro-oxidant molecules released during malaria contribute in pathological complications of cerebral malaria, vascular complication and immune-depression following multiple mechanisms, but still it is not conclusive [28], [29], [30].…”

Section: Introductionmentioning

confidence: 99%

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Phagocytic Uptake of Oxidized Heme Polymer Is Highly Cytotoxic to Macrophages

Deshmukh

Trivedi

2014

PLoS ONE

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Apoptosis in macrophages is responsible for immune-depression and pathological effects during malaria. Phagocytosis of PRBC causes induction of apoptosis in macrophages through release of cytosolic factors from infected cells. Heme polymer or β-hematin causes dose-dependent death of macrophages with LC50 of 132 µg/ml and 182 µg/ml respectively. The toxicity of hemin or heme polymer was amplified several folds in the presence of non-toxic concentration of methemoglobin. β-hematin uptake in macrophage through phagocytosis is crucial for enhanced toxicological effects in the presence of methemoglobin. Higher accumulation of β-hematin is observed in macrophages treated with β-hematin along with methemoglobin. Light and scanning electron microscopic observations further confirm accumulation of β-hematin with cellular toxicity. Toxicological potentiation of pro-oxidant molecules toward macrophages depends on generation of H2O2 and independent to release of free iron from pro-oxidant molecules. Methemoglobin oxidizes β-hematin to form oxidized β-hematin (βH*) through single electron transfer mechanism. Pre-treatment of reaction mixture with spin-trap Phenyl-N-t-butyl-nitrone dose-dependently reverses the β-hematin toxicity, indicates crucial role of βH* generation with the toxicological potentiation. Acridine orange/ethidium bromide staining and DNA fragmentation analysis indicate that macrophage follows an oxidative stress dependent apoptotic pathway to cause death. In summary, current work highlights mutual co-operation between methemoglobin and different pro-oxidant molecules to enhance toxicity towards macrophages. Hence, methemoglobin peroxidase activity can be probed for subduing cellular toxicity of pro-oxidant molecules and it may in-turn make up for host immune response against the malaria parasite.

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Section: Resultsmentioning

confidence: 98%

Section: Discussionmentioning

confidence: 99%

Section: Methodsmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

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Phagocytic Uptake of Oxidized Heme Polymer Is Highly Cytotoxic to Macrophages

Deshmukh

Trivedi

2014

PLoS ONE

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“…The results demonstrate an increase in deoxyhemoglobin and methemoglobin concentration by 1.34 and 2.81 respectively, and a decrease in oxyhemoglobin concentration by 1.07. The formation of methemoglobin has important roles in inflammation and vascular regulation [10], [26]. Spectrophotometer measurements demonstrate changes in the optical properties because of adding ceramide ( Figure 3).…”

Section: Resultsmentioning

confidence: 99%

Photoacoustic spectral analysis to sense programmed erythrocyte cell death (eryptosis) for monitoring cancer response to treatment

2016

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Many types of cancer therapies target the tumor microenvironment, causing biochemical and morphological changes in tissues. In therapies using ultrasound activated microbubbles, vascular collapse is typically reported. Red blood cells (RBCs) that leak out of the vasculature become exposed to the ceramide that is released from damaged endothelial cells. Ceramide can induce programmed cell death in RBCs (eryptosis), and is characterized by cell shrinkage, membrane blebbing and scrambling. Since the effect of eryptotic cells on generated photoacoustics (PA) signals has not been reported, we investigated the potential PA may have for cancer treatment monitoring by using PA spectral analysis to sense eryptosis. To induce eryptosis, C2-ceramide was added to RBC suspensions and that were incubated for 24 hours at 37 o C. A control and ceramide-induced sample was imaged in a vessel phantom using a high frequency PA system (VevoLAZR, 10 -45 MHz bandwidth) irradiated with multiple wavelengths ranging from 680 to 900 nm. PA spectral parameters were measured and linked to changes in RBCs as it underwent eryptosis. These samples were examined using optical microscopy, a blood gas analyzer and an integrating sphere setup to measure optical properties (wavelengths 600 -900 nm). The results of the experiment demonstrate how PA spectral analysis can be used to identify eryptosis at a depth of more than 1 cm into the phantom using ultrasound derived the y-intercept and mid bandfit (MBF) parameters at optical wavelengths of 800 -900 nm. These parameters were correlated to the morphological and biochemical changes that eryptotic RBCs display. The results establish the potential of PA in cancer treatment monitoring through sensing treatment induced eryptosis.

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Optical and photoacoustic radiofrequency spectroscopic analysis for detecting red blood cell death

Fadhel

Hysi

Strohm

et al. 2019

Journal of Biophotonics

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Under stress, red blood cells (RBCs) undergo programmed cell death (eryptosis). One of the signaling molecules for eryptosis, sphingomyelinase (SMase), plays an important role in monitoring the efficacy of vascular targeted cancer therapy. The high optical absorption of erythrocytes coupled with the changes of eryptotic RBCs makes RBCs ideal targets for the photoacoustic (PA) detection and characterization of vascular treatments. In this work, experiments characterizing eryptosis were performed: PA detection of high frequencies (>100 MHz) that enabled analysis at the single‐cell level and of low frequencies (21 MHz) that enabled analysis at the RBC ensemble level. Ultrasound spectral analysis was performed on control and SMase‐treated RBCs. Spectral unmixing was applied to quantify methemoglobin production as a by‐product of RBC death. Validation was performed using a blood gas analyzer and optical spectrometry. Our results indicate that PA radiofrequency spectra could be used to differentiate the biochemically induced morphological changes as RBCs lose their native biconcave shape, and release hemoglobin into the surroundings. Spectral unmixing revealed a 7% increase in methemoglobin content for SMase‐treated samples due to the oxidative stress on the RBCs. These findings suggest that PA spectral analysis of RBC death can potentially serve as a biomarker of the efficacy of vascular targeted cancer therapies.

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Extracellular methemoglobin primes red blood cell aggregation in malaria: An in vitro mechanistic study

Cited by 23 publications

References 45 publications

Phagocytic Uptake of Oxidized Heme Polymer Is Highly Cytotoxic to Macrophages

Phagocytic Uptake of Oxidized Heme Polymer Is Highly Cytotoxic to Macrophages

Photoacoustic spectral analysis to sense programmed erythrocyte cell death (eryptosis) for monitoring cancer response to treatment

Optical and photoacoustic radiofrequency spectroscopic analysis for detecting red blood cell death

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