Binding Of Mb Research Articles

A-015 Changes of sST2 and NT-proBNP levels predict early cardiovascular toxicity in breast cancer patients treated with anthracycline-containing chemotherapy

Abstract Background Cardiovascular biomarkers play a crucial role in monitoring the risk for cancer therapy-related cardiovascular toxicity, but their changes and values at the early stage of chemotherapy are still inadequate, especially for some new biomarkers.The purpose of this study was to examine changes in cardiovascular biomarkers and evaluate the prognostic values for cardiovascular toxicity in patients with breast cancer treated with anthracycline-containing chemotherapy. Methods In a prospective cohort of 73 patients treated with anthracycline-containing chemotherapy, soluble ST2 (sST2), high-sensitivity cardiac troponin T (hs-cTnT), N-terminal pro-B-type natriuretic peptide (NT-proBNP), Myoglobin (Myo), Creatine kinase isoenzyme MB (CK-MB) and heart-fatty acid binding protein (H-FABP) were measured at baseline, during every chemotherapy cycle (C1-C6, about every month).Cardiovascular toxicity was defined as cancer therapy-related arrhythmias, which was differentiated into bradycardia,tachycardia or atrial fibrillation. According to whether arrhythmia occurred, patients were divided into two groups (healthy group and arrhythmias group), and the basic clinical characteristics were collected and compared among the two groups. Logistic regression analyses and receiver operating characteristic (ROC) curves were conducted to investigate the relationship between the changes in biomarkers and anthracycline-induced cardiotoxicity. Results sST2 levels increased significantly from baseline to C1 (P<0.01). NT-proBNP decreased from baseline to C1, C5 (P< 0.01). Logistic regression analysis showed a greater risk of cardiovascular toxicity was associated with greater interval increase in sST2 from baseline to C1(ΔsST2) (OR:1.27; 95% CI:1.03 to1.56; P= 0.024) and interval decline in NT-proBNP from baseline to C1(ΔNT-proBNP) was associated with cardiovascular toxicity (OR: 0.83; 95% CI: 0.70 to 0.98; P= 0.029). ROC curves showed that ΔsST2, ΔNT-proBNP and ΔsST2+ΔNT-proBNP had good predictive value for cardiovascular toxicity (areas under the curves were 0.631, 0.633, and 0.735, respectively, P<0.05). Conclusions Early changes in sST2 and NT-proBNP levels offer additive information in early cardiovascular toxicity risk prediction in patients with breast cancer who receive anthracycline-containing chemotherapy.

Contrast-enhanced ultrasound molecular imaging of activated platelets in the progression of atherosclerosis using microbubbles bearing the von Willebrand factor A1 domain.

Platelet-endothelial interactions have been linked to increased inflammatory activation and a prothrombotic state in atherosclerosis. The interaction between von Willebrand factor (vWF)-A1 domain and platelet glycoprotein (GP) Ib/IX plays a significant role in mediating the adhesion of platelets to the injured endothelium. In the present study, contrast-enhanced ultrasound (CEU) molecular imaging with microbubbles bearing the vWF-A1 domain was performed to non-invasively monitor activated platelets on the vascular endothelium in the procession of atherosclerosis. A targeted CEU contrast agent was prepared by attaching the vWF-A1 domain to the shell of microbubbles (MbA1). Rat isotype control antibody was used to produce control (Mbctrl) microbubbles. The binding of MbA1 and Mbctrl to activated platelets was assessed in in vitro flow chamber experiments. Apolipoprotein E (ApoE-/-) deficient mice were studied as a model of atherosclerosis. At 8, 16 and 32 weeks of age, CEU molecular imaging of the proximal aorta with MbA1 and Mbctrl was performed and the imaging signals from microbubbles were quantified. Atherosclerotic lesion severity and platelets on the endothelial surface were assessed by histology and immunohistochemistry. In in vitro flow chamber studies, attachment of MbA1 to activated platelets on culture dishes was significantly greater than that of Mbctrl across a range of shear stresses (P<0.05). The attachment of Mbctrl was sparse and not related to the aggregated platelets. As lesion development progressed in the ApoE-/- mice, molecular imaging of activated platelets demonstrated selective signal enhancement of MbA1 (P<0.05 vs. Mbctrl) at all ages. Selective signal enhancement from MbA1 increased from 8 to 32 weeks of age. Immunohistochemistry for GPIIb revealed the presence of platelets on the endothelial cell surface in each group of ApoE-/- mice and that the degree of platelet deposits was age-dependent. The results of the present study indicated that non-invasive CEU molecular imaging with targeted microbubbles bearing the vWF-A1 domain could not only detect activated platelets on the vascular endothelium but also indicate lesion severity in atherosclerosis.

Turning date palm waste into carbon nanodots and nano zerovalent iron composites for excellent removal of methylthioninium chloride from water

Novel carbon nanodots (nCD-DBC) and nano zero-valent iron composites (nZVI-DBC) were synthesized using date palm waste-derived biochar (DBC). The synthesized materials were analyzed for chemical and structural composition by using FTIR, SEM, XRD, and TGA, and evaluated for their methylthioninium chloride dye (MB) removal efficiency from contaminated aqueous solutions. pH 7.0 was found optimum for the highest MB removal in sorption batch studies. Kinetics sorption of MB onto the sorbents was best described by pseudo-second-order (R2 = 0.93–0.99) and Elovich models (R2 = 0.86–0.97) implying that sorption was being controlled by chemisorption. Langmuir model predicted maximum sorption capacities for nCD-DBC, nZVI-DBC, and DBC were 1558.66, 1182.90, and 851.67 mg g−1, respectively, which correlated with the results of kinetics sorption. Likewise, nCD-DBC yielded the highest partition coefficient (7067 mL g−1), followed by nZVI-DBC (1460 mL g−1), and DBC (930 mL g−1). Post-sorption XRD, FTIR, and SEM analyses depicted the binding of MB onto the sorbents. It was suggested that electrostatic interactions, π–π electron donor-accepter interactions, degradation, and diffusion were responsible for MB removal by the synthesized materials. Therefore, the nCD-DBC, nZVI-DBC, and DBC can potentially be used for scavenging MB dye from contaminated aqueous solutions.

Label-free aptasensor for the detection of cardiac biomarker myoglobin based on gold nanoparticles decorated boron nitride nanosheets

A novel electrochemical aptasensor based on gold nanoparticles decorated on boron nitride nanosheets (AuNPs/BNNSs) for the sensitive and selective detection of myoglobin (Mb) is reported. BNNSs were chemically synthesized by a low-cost and simple hydrothermal method. They were deposited onto the fluorine-doped tin oxide (FTO) electrode by a spin-coating method. Subsequently, AuNPs were chemically deposited onto the BNNS/FTO electrode by a seed-mediated chemical reduction method, with an average particle size of approximately 10 nm. The AuNPs/BNNSs/FTO electrode was used as a transducer to immobilize a thiol-functionalized DNA aptamer (Apt) via the covalent interaction of Au–S for the specific binding of Mb. [Fe(CN)6]3-/4- was used as a redox probe to monitor the oxidation current variation upon the binding of Mb with varying concentrations onto the sensor surface. The Apt/AuNPs/BNNSs/FTO sensor shows a high signal response for Mb with a detection limit of 34.6 ng/mL and a dynamic response range of 0.1–100 µg/mL. It is a promising candidate for point-of-care diagnosis in real samples. This strategy could make possible the application of other 2D materials with wide bandgaps for the development of biosensors.

Insonation of Systemically Delivered Cisplatin-Loaded Microbubbles Significantly Attenuates Nephrotoxicity of Chemotherapy in Experimental Models of Head and Neck Cancer.

The use of cisplatin (CDDP), the most common chemotherapy drug for head and neck cancer, is limited by its undesirable side effects, especially nephrotoxicity. We investigated ultrasound microbubbles (USMB) as a tool to increase the local intra-tumoral CDDP level while decreasing systemic CDDP cytotoxicity. We allowed CDDP to interact with human serum albumin and then sonicated the resulting CDDP‒albumin complex to generate CDDP-loaded MBs (CDDP-MBs). We then established a head-and-neck tumor-bearing mouse model by implanting FaDu-fLuc/GFP cells into severe combined immunodeficiency mice and used IVIS® bioluminescence imaging to determine the tumor xenograft formation and size. Twice weekly (until Day 33), we administered CDDP only, CDDP + MBs + US, CDDP-MBs, or CDDP-MBs + US intravenously by tail-vein injection. The US treatment was administered at the tumor site immediately after injection. The in vivo systemic distribution of CDDP indicated that the kidney was the most vulnerable organ, followed by the liver, and then the inner ear. However, CDDP uptake into the kidney and liver was significantly decreased in both the CDDP-MBs and CDDP-MBs + US groups, suggesting that MB binding significantly reduced the systemic toxicity of CDDP. The CDDP-MBs + US treatment reduced the tumor size as effectively as conventional CDDP-only chemotherapy. Therefore, the combination of CDDP-MBs with ultrasound is effective and significantly attenuates CDDP-associated nephrotoxicity, indicating a promising clinical potential for this approach.

Adsorptive performance of MOF nanocomposite for methylene blue and malachite green dyes: Kinetics, isotherm and mechanism

In the present study, Fe3O4@AMCA-MIL-53(Al) nanocomposite was utilized for the adsorptive removal of highly toxic MB and MG dyes from aqueous environment. The batch adsorption tests were performed at different contact time, pH, Fe3O4@AMCA-MIL-53(Al) dose, initial concentration of dyes and temperature. The maximum adsorption capacity of MB and MG dyes onto of Fe3O4@AMCA-MIL-53(Al) using Langmuir equation was 1.02 and 0.90 m mol/g, respectively. The isotherm and kinetic studies revealed that adsorption data were well fitted to Langmuir isotherm and pseudo-first-order kinetics models. Various thermodynamic parameters were also calculated and interpreted. The positive and negative values of ΔH° and ΔG° indicated that the adsorption was endothermic and spontaneous, respectively. The adsorptive binding of MB and MG on Fe3O4@AMCA-MIL53(Al) nanocomposite was directed by carboxylate and amide groups through electrostatic interaction, π−π interaction and hydrogen bonding. The desorption of both dyes from Fe3O4@AMCA-MIL-53(Al) was also performed using mixed solution of 0.01 M HCl/ethanol. Thus, we conclude that the Fe3O4@AMCA-MIL-53(Al) was an outstanding material for the removal of dyes from aqueous environment.

Abstract 357: Rapid harvesting of highly concentrated circulating tumor cells from bulk human blood with buoyant immuno-microbubbles

Abstract Circulating tumor cells (CTCs) are the extremely rare cells shed from primary tumor during metastatic disease. A technology to quickly and inexpensively isolate viable CTCs will enable widespread clinical utilization of CTCs for cancer prognosis and drug testing. The lipid-shelled microbubble is a biocompatible material that has been used as ultrasound contrast media. We have previously shown that MB is a promising vehicle to isolate rare cells from biological fluids by flotation. Here we investigated the role of lipid composition, MB/cell ratio, and type of tumor surface marker [epithelial cell adhesion molecule (EpCAM), receptor tyrosine-protein kinase erbB-2 or human epidermal growth factor receptor 2 (HER2/neu) and epithelial growth factor receptor (EGFR)] on MB stability and binding efficiency to breast cancer cells. Optimized MBs coated with anti-EpCAM antibody showed 95% binding efficiency to human breast carcinoma SKBR-3 cells. Cells were able to proliferate in culture with MBs attached to them. We further developed an inverted setup in order to harvest (skim) highly concentrated tumor cells in a microliter volume on a nitrocellulose membrane. Using vacuum-assisted harvesting we demonstrated a high efficiency (∼80%) of isolation of rare tumor cells spiked in 7 ml blood by anti-EpCAM MBs. A blinded study using anti-EpCAM MBs detected epithelial CTCs in samples of metastatic breast and ovarian cancers with brain metastases, but not in samples from non-epithelial brain malignancies, including primary glioblastoma multiforme, hemangioblastoma and metastatic melanoma. The simple and robust MB approach can be developed into an inexpensive tool for batch CTC isolation in basic research and clinical applications. Citation Format: Guankui Wang, Sandeep Pingle, Santosh Kesari, Yu-Tsueng Liu, Dmitri Simberg. Rapid harvesting of highly concentrated circulating tumor cells from bulk human blood with buoyant immuno-microbubbles. [abstract]. In: Proceedings of the 107th Annual Meeting of the American Association for Cancer Research; 2016 Apr 16-20; New Orleans, LA. Philadelphia (PA): AACR; Cancer Res 2016;76(14 Suppl):Abstract nr 357.

Behavior of Ethidium Bromide-Hoechst 33258-DNA and Ethidium Bromide-Methylene Blue-DNA Triple Systems by means of UV Melting

The study of EtBr and H33258 interaction as well as EtBr and MB interaction with DNA has been carried out. It was revealed that, at joint interaction, the effect of two ligands on the change of melting thermodynamic parameters of EtBr-DNA-H33258 or EtBr-DNA-MB complexes is not an addition of separate interaction influences. It was shown that, at joint, binding of EtBr and MB with DNA competition occurs, while in the case of EtBr and H33258, the mutual strengthening of stabilizing effect of each of them on DNA double-stranded structure mainly takes place.

Protein electronic conductors: hemin-substrate bonding dictates transport mechanism and efficiency across myoglobin.

Electron transport (ETp) across met-myoglobin (m-Mb), as measured in a solid-state-like configuration between two electronic contacts, increases by up to 20 fold if Mb is covalently bound to one of the contacts, a Si electrode, in an oriented manner by its hemin (ferric) group, rather than in a non-oriented manner. Oriented binding of Mb is achieved by covalently binding hemin molecules to form a monolayer on the Si electrode, followed by reconstitution with apo-Mb. We found that the ETp temperature dependence (>120 K) of non-oriented m-Mb virtually disappears when bound in an oriented manner by the hemin group. Our results highlight that combining direct chemical coupling of the protein to one of the electrodes with uniform protein orientation strongly improves the efficiency of ET across the protein. We hypothesize that the behavior of reconstituted m-Mb is due to both strong protein-substrate electronic coupling (which is likely greater than in non-oriented m-Mb) and direct access to a highly efficient transport path provided by the hemin group in this configuration.

Protein Electronic Conductors: Hemin–Substrate Bonding Dictates Transport Mechanism and Efficiency across Myoglobin

AbstractElectron transport (ETp) across met‐myoglobin (m‐Mb), as measured in a solid‐state‐like configuration between two electronic contacts, increases by up to 20 fold if Mb is covalently bound to one of the contacts, a Si electrode, in an oriented manner by its hemin (ferric) group, rather than in a non‐oriented manner. Oriented binding of Mb is achieved by covalently binding hemin molecules to form a monolayer on the Si electrode, followed by reconstitution with apo‐Mb. We found that the ETp temperature dependence (&gt;120 K) of non‐oriented m‐Mb virtually disappears when bound in an oriented manner by the hemin group. Our results highlight that combining direct chemical coupling of the protein to one of the electrodes with uniform protein orientation strongly improves the efficiency of ET across the protein. We hypothesize that the behavior of reconstituted m‐Mb is due to both strong protein–substrate electronic coupling (which is likely greater than in non‐oriented m‐Mb) and direct access to a highly efficient transport path provided by the hemin group in this configuration.

Noninvasive molecular ultrasound monitoring of vessel healing after intravascular surgical procedures in a preclinical setup.

Cardiovascular interventions induce damage to the vessel wall making antithrombotic therapy inevitable until complete endothelial recovery. Without a method to accurately determine the endothelial status, many patients undergo prolonged anticoagulation therapy, denying them any invasive medical procedures, such as surgical operations and dental interventions. Therefore, we aim to introduce molecular ultrasound imaging of the vascular cell adhesion molecule (VCAM)-1 using targeted poly-n-butylcyanoacrylate microbubbles (MB(VCAM-1)) as an easy accessible method to monitor accurately the reendothelialization of vessels. ApoE(-/-) mice were fed with an atherogenic diet for 1 and 12 weeks and subsequently, endothelial denudation was performed in the carotid arteries using a guidewire. Molecular ultrasound imaging was performed at different time points after denudation (1, 3, 7, and 14 days). An increased MB(VCAM-1) binding after 1 day, a peak after 3 days, and a decrease after 7 days was found. After 12 weeks of diet, MB(VCAM-1) binding also peaked after 3 days but remained high until 7 days, indicating a delay in endothelial recovery. Two-photon laser scanning microscopy imaging of double fluorescence staining confirmed the exposure of VCAM-1 on the superficial layer after arterial injury only during the healing phase. After complete reendothelialization, VCAM-1 expression persisted in the subendothelial layer but was not reachable for the MBV(CAM-1) anymore. Molecular ultrasound imaging with MB(VCAM-1) is promising to assess vascular damage and to monitor endothelial recovery after arterial interventions. Thus, it may become an important diagnostic tool supporting the development of adequate therapeutic strategies to personalize anticoagulant and anti-inflammatory therapy after cardiovascular intervention.

DNA Electrochemical Sensor for Detection of PRSS1 Point Mutation Based on Restriction Endonuclease Technique

To construct a restriction endonuclease based biosensor technology for PRSS1 genotyping. We designed a thiol-modified hairpin probe where the neck has EcoRI endonuclease recognition sites according to the PRSS1 gene c.410 C>T (p.T137 M) mutation and it was fixed on the gold electrode. Different charge generated by the binding of MB to phosphate groups of DNA before and after hybridization was used for distinguishing the different genotypes and quantity. This showed that the novel sensor can better distinguish the complementary sequence, single-base mismatches, and completely noncomplementary sequences, and the linear range for the logarithm was Y = –0.0242 X + 0.1574, R = 0.9912(Y = current, X = log target DNA concentration); the detection limit for DNA detection is estimated to be 50 fM.

Synergetic gating of metal-latching ligands and metal-chelating proteins for mesoporous silica nanovehicles to enhance delivery efficiency.

Stimuli-responsive drug delivery systems are highly desirable for improved therapeutic efficacy and minimized adverse effects of drugs. Mesoporous silica nanoparticles (MSNs) functionalized with pentadentate ligands, N-(3-trimethoxysilylpropyl)ethylenediamine triacetate (TSP-DATA), in the presence of metal ions with and without myoglobin (Mb)-containing surface-accessible histidine residues, were constructed for pH-triggered controlled release. The DATA ligands immobilized on the MSN pore outlets could encapsulate cargo within the pores by metal latching across pore openings, and release efficiency increased with the increase of surface density of the DATA ligands. The release efficiencies for the metal-chelating protein nanogates, through multiple-site binding of Mb with the metal-chelating ligands, were higher than those for the metal-latching ligand nanogates but were almost independent of surface density of the ligands investigated. Both the metal-latching ligands and the metal-chelating proteins played a synergetic role in gating MSNs for high-loading drug delivery and stimuli-responsive controlled release. The constructed Mb-Cu(2+)-gated MSN delivery system has promising applications in targeted drug therapy of tumors.

A rapid and sensitive method for the determination of dibutyl phthalate in wine by flow-injection chemiluminescence analysis

A sensitive method for the determination of picogram level dibutyl phthalate (DBP) in wine by flow-injection chemiluminescence (FI–CL) analysis is presented for the first time, which was based on the quenching effect of DBP on the luminol–myoglobin (Mb) CL system. The decrement of CL intensity was linearly proportional to the logarithm of DBP concentration in the range of 0.1–100pgmL−1 with the detection limit of 0.03pgmL−1 (3σ). At a flow rate of 2.0mLmin−1, a complete determination of DBP including sampling and washing could be accomplished in 0.5min, giving the maximum sample throughput of 120h−1. The proposed method was successfully applied to the determination of DBP in wine, human serum and urine samples with the relative standard deviations (RSDs) of less than 3.0% (n=5). The molecule docking results showed that DBP interacted with the amino acid residues near the heme moiety of Mb. The possible CL mechanism of luminol–Mb–DBP reaction should be that the binding of Mb with DBP forming a 1:1 complex (binding constant K=1.55×104Lmol−1) led to the conformational change of Mb and resulted in the quenching of CL intensity.

Rebinding Dynamics of CO Following Photodissociation of 4.0 M Guanidine HCl-Denatured Carbonmonoxyhemoglobin

13 CO. Geminate rebinding of CO to the denatured Hb was accelerated more than 1000 times, suggesting that the native structure of the Hb is required to suppress efficient geminate rebinding of CO, as is the case in Mb. The geminate yield and rate for CO rebinding are almost the same in both the denatured Hb and Mb. Similarity in the equilibrium spectrum and rebinding dynamics of CO indicates that the state of the denatured Hb is very similar to that of the denatured Mb. In the denatured Hb, quaternary contact of the protein is likely severed, with the denatured protein existing as an independent subunit much like Mb. Mb, an oxygen storage protein, is a monomeric protein that contains a single heme group as the active ligand-binding site of the protein. Hb, an oxygen transport protein, is a tetrameric protein, each unit of which contains a heme group, with the global tertiary structure of each unit similar to that of Mb. Hb exhibits cooperative binding of ligands essential to efficient transport of O2. 2 The cooperativity of Hb is accompanied by a substantial change in its quaternary structure. Whereas the peptide sequence of Mb is different from that of Hb, amino acid residues near the heme in both proteins are highly conserved. The residues surrounding the bound ligand are invariant in both proteins, except for isoleucine 107 in Mb that is replaced with leucine 107 in Hb. The rebinding of CO to Mb and Hb after photolysis are most often used to study the relationships between protein structure, function, and dynamics. Though the rebinding dynamics of the ligands to both proteins are similar in general, they are different in detail. Whereas cooperative binding of the ligand to Hb makes its behavior difficult to compare directly to the binding of Mb, the difference is often attributed to arise from the quaternary structure of Hb. The dynamics and structural characterization of denatured proteins can be useful in understanding protein folding as well as various biological phenomena such as regulation of the cellular activity and onset of neurodegenerative disease. 3

Interaction of Fish Myoglobin and Myofibrillar Proteins

Interactions between fish myoglobin (Mb) and myofibrillar proteins were investigated in a Mb-natural actomyosin (NAM) model at 4 degrees C. Increases in metmyoglobin (MetMb) formation and the relative content of bound Mb were observed, as were decreases in whiteness and Ca(2+)-ATPase activity (P < 0.05). During the first 6 h of incubation, Mb bound preferably to myosin at domains other than the head portion, as evidenced by measurable ATPase activity. The potential binding of Mb to myosin heads occurred after 24-h incubation as evidenced by the marked decrease in Ca(2+)-ATPase activity of the NAM-Mb mixture when compared to that of NAM alone (P < 0.05). The interaction between fish Mb and myofibrillar proteins was more pronounced with increased storage time; formation of high-molecular-weight aggregates (> 206 kDa) also increased with time. Electrophoretic study revealed that disulfide bonds were not involved in Mb-NAM interactions.

A Controllable Solid-State Ru(bpy)32+ Electrochemiluminescence Film Based on Conformation Change of Ferrocene-Labeled DNA Molecular Beacon

A controllable solid-state electrochemiluminescence (ECL) film based on efficient and stable quenching of ECL of ruthenium(II) tris-(bipyridine) (Ru(bpy)32+) by oxidizing ferrocene (Fc) at the electrode is developed. The ECL intensity is correlated to the distance which is controlled by the conformation of the ferrocene-labeled DNA molecular beacon (Fc-MB) between the Fc and Ru(bpy)32+ immobilized on the electrode. The conformation adjustment is conducted via complementary DNA hybridizing with the bases in the loop of the Fc-MB and changing the temperature of the Fc-MB and the resultant double-stranded DNA (dsDNA). Those events all result in change of the ECL intensity. With such characteristics, the solid-state Ru(bpy)32+-ECL film has the potential to be applied to reagentless DNA ECL biosensors and to calculate thermodynamic parameters of equilibrium constants of MB binding and the stem-loop formation.

Effects of Ionic Surfactants and Cyclodextrins on Hydride-Transfer Reaction of 1-Benzyl-1,4-dihydronicotinamide with Methylene Blue

Abstract The kinetics of the hydride-transfer reaction between methylene blue (MB+) and 1-benzyl-1,4-dihydronictinamide (BNAH) were studied in media containing cyclodextrins (β- and γ-CD) and surfactants (sodium dodecyl sulfate (SDS), dodecyltrimethylammonium bromide, tetradecyltrimethylammonium bromide, and hexadecyltrimethylammonium bromide). Cationic surfactants decreased the apparent first-order rate constant (kobsd) above the cmc, while SDS increased kobsd just above the cmc and then decreased kobsd with increasing surfactant concentration. This behavior for cationic surfactants was typical of micellar effects due to a separation of the reactants by the micelles. BNAH associated with micelles, whereas MB+ ions were repelled from the cationic interface of the micelles. Binding of BNAH and MB+ to the same SDS micelle enhanced the reaction, but dilution of reagents within the micellar interface with the increase in [SDS] caused a decrease in kobsd. In β-CD–cationic surfactant mixtures, the results were interpreted in terms of the model which takes into account the formation of CD–BNAH, CD–MB+, and CD–surfactant complexes and the association of BNAH with micelles. The decrease in kobsd with increasing surfactant concentration observed in γ-CD–cationic surfactant mixtures can be explained by the decrease in the concentration of free γ-CD by the formation of 1:1 and 2:1 complexes of surfactant monomer with γ-CD.

Resensitization of breast cancer cells to anoikis by Tropomyosin-1: role of Rho kinase-dependent cytoskeleton and adhesion

Two most common properties of malignant cells are the presence of aberrant actin cytoskeleton and resistance to anoikis. Suppression of several key cytoskeletal proteins, including tropomyosin-1 (TM1), during neoplastic transformation is hypothesized to contribute to the altered cytoskeleton and neoplastic phenotype. Using TM1 as a paradigm, we have shown that cytoskeletal proteins induce anoikis in breast cancer (MCF-7 and MDA MB 231) cells. Here, we have tested the hypothesis that TM1-mediated cytoskeletal changes regulate integrin activity and the sensitivity to anoikis. TM1 expression in MDA MB 231 cells promotes the assembly of stress fibers, induces rapid anoikis via caspase-dependent pathways involving the release of cytochrome c. Further, TM1 inhibits binding of MDA MB 231 cells to collagen I, but promotes adhesion to laminin. Inhibition of Rho kinase disrupts TM1-mediated cytoskeletal reorganization and adhesion to the extracellular matrix components, whereas the parental cells attach to collagen I, spread and form extensive actin meshwork in the presence of Rho kinase inhibitor, underscoring the differences in parental and TM1-transduced breast cancer cells. Further, treatment with the cytoskeletal disrupting drugs rescues the cells from TM1-induced anoikis. These new findings demonstrate that the aberrant cytoskeleton contributes to neoplastic transformation by conferring resistance to anoikis. Restoration of stress fiber network through enhanced expression of key cytoskeletal proteins may modulate the activity of focal adhesions and sensitize the neoplastic cells to anoikis.

Binding, aggregation and photochemical properties of methylene blue in mitochondrial suspensions

AbstractMethylene Blue (MB) has well‐established photochemical properties and has been used in a variety of photochemical applications including photodynamic therapy. Despite the fact that most of MB's cytotoxic effects in cells are attributed to mitochondrial damage, the interactions of this dye with mitochondria and the consequent effects on photochemical properties have not yet been fully determined. We monitored MB binding, aggregation and its ability to release singlet oxygen (1O2) on irradiation when interacting with mitochondrial suspensions. MB actively binds to mitochondria and enters the matrix in a manner stimulated by the mitochondrial proton potential and by the increase in mitochondrial concentrations. The greater accumulation of MB in mitochondria with elevated proton potentials or those treated with high concentrations of MB results in the formation of MB dimers, previously shown to be less effective generators of 1O2. Accumulation of MB within mitochondria with high membrane potentials also results in the reduction of MB to the photochemically inactive leuco‐MB. Indeed, irradiation of mitochondria with high proton potentials in the presence of MB results in the generation of approximately half the quantity of 1O2 compared with 1O2 generated in mitochondria with low proton potentials. These differences in photochemical properties should influence the cytotoxic effects of photodynamic treatment in the presence of MB.