Dual pH Research Articles

Effects of Resveratrol on Mitochondrial Autophagy in Doxorubicin Treated Cardiac Myoblast Cells

Background and ObjectiveDoxorubicin (DOX) is a widely used chemotherapeutic agent with dose‐dependent cardiotoxic effects. It is of great clinical significance to identify a cytoprotective agent which can attenuate DOX cardiotoxicity without affecting its anti‐tumor efficacy. Resveratrol (Rev) is a plant‐derived polyphenol shown to exert cardioprotective effects in DOX‐treated cardiomyocytes, but its mechanisms of cardioprotection remain partially understood. We tested the hypothesis that Rev protects against DOX‐induced cardiomyocyte injury by inhibiting excessive degradation of mitochondria through the autophagy‐lysosome pathway (mitophagy).
MethodsH9C2 cardiac myoblasts were cultured under four conditions: 0.5 uM DOX, 5 uM Rev, 5 uM Rev + 0.5 uM DOX, and Ethanol (control). Propidium Iodide (PI) staining was used to assess necrotic cell death. Western Blotting was used to measure PARP cleavage, an apoptotic marker. Mitophagy was assessed via cellular infection with Adenovirus encoding MitoRosella, a novel dual fluorescent pH sensitive mitophagy reporter.
ResultsDOX induced cardiomyocyte death as measured by PI positive cell and PARP cleavage, which was associated with increased mitophagy as assessed by the MitoRosella reporter. Pre‐treatment of cells with Rev not only attenuated DOX‐induced cell death but also inhibited DOX‐induced excessive mitophagy (Fig.1 C‐D). Rev alone had no effect on mitophagy (Fig.1 A‐B).imageConclusionThe cardioprotective effect of Resveratrol is associated with a significant reduction in DOX‐induced mitophagy in cardiomyocytes, suggesting the excessive destruction of mitochondria as a potential cardiotoxic mechanism of DOX.

Dual pH- and temperature-responsive protein nanoparticles

Multiply responsive protein nanoparticles are interesting for a variety of applications. Herein, we describe the synthesis of a vault nanoparticle that responds to both temperature and pH. Specifically, poly(N-isopropylacrylamide-co-acrylic acid) with a pyridyl disulfide end group was prepared by reversible addition-fragmentation chain transfer (RAFT) polymerization. The polymer had a lower critical solution temperature (LCST) of 31.9°C at pH 5, 44.0°C at pH 6 and above 60°C at pH 7. The polymer was conjugated to human major vault protein (hMVP), and the resulting nanoparticle was analyzed by UV–Vis, dynamic light scattering (DLS) and electron microscopy. The data demonstrated that the poly(N-isopropylacrylamide-co-acrylic acid)-vault conjugate did not respond to temperatures below 60°C at pH 7, while the nanoparticles reversibly aggregated at pH 6. Furthermore, it was shown that the vault nanoparticle structure remained intact for at least three heat and cooling cycles. Thus, these dually responsive nanoparticles may serve as a platform for drug delivery and other applications.

Comparison of Type 1 D-3-phosphoglycerate dehydrogenases reveals unique regulation in pathogenic Mycobacteria

D-3-phosphoglycerate dehydrogenases (PGDH) from all organisms catalyze the conversion of D-3-phosphoglycerate to phosphohydroxypyruvate as the first step in the biosynthesis of l-serine. This investigation compares the properties of Type 1 PGDHs from seven different species and demonstrates that conserved residues in the ACT and ASB domains of some allow l-serine to act as a feedback inhibitor at low micromolar concentrations. In addition, the serine sensitivity is dependent on the presence of phosphate ions. These residues are most highly conserved among PGDHs from the actinomycetales family, but only certain pathogenic mycobacteria appear to have the full complement of residues required for high sensitivity to serine. These basic residues are also responsible for the presence of dual pH optima in the acidic region that is also phosphate dependent. Analytical ultracentrifugation analysis demonstrates that the dual pH optima do not require changes in oligomeric state. This study also demonstrates that substrate inhibition is a common feature of Type 1 PGDHs and that it is suppressed by phosphate, indicating that phosphate likely interacts at both the catalytic and regulatory sites. The unique features resulting from the complement of basic residues conserved in pathogenic mycobacteria may impart important metabolic advantages to these organisms.

Production and characterization of thermostable alkaline protease of Bacillus subtilis (ATCC 6633) from optimized solid-state fermentation.

Proteases are the most important group of enzymes utilized commercially in various arenas of industries, such as food, detergent, leather, dairy, pharmaceutical, diagnostics, and waste management, accounting for nearly 20% of the world enzyme market. Microorganisms of specially Bacillus genera serve as a vast repository of diverse set of industrially important enzymes and utilized for the large-scale enzyme production using a fermentation technology. Approximately 30%-40% of the cost of industrial enzymes originates from the cost of the growth medium. This study is attempted to produce protease from Bacillus subtilis (ATCC 6633) after optimization of various process parameters with the aid of solid-state fermentation using a cheap nutrient source such as wheat bran. B. subtilis (ATCC 6633) produces proteases of molecular weight 36 and 20 kDa, respectively, in the fermented medium as evident from SDS zymogram. Alkaline protease activity has been detected with optimum temperature at 50°C and is insensitive to ethylenediaminetetraacetic acid. This thermostable alkaline protease exhibits dual pH optimum at 7 and 10 with moderate pH stability at alkaline pH range. It preserves its activity in the presence of detergent such as SDS, Tween 20, and Triton X-100 and may be considered as an effective additive to detergent formulation with some industrial importance.

PH and glucose responsive nanofibers for the reversible capture and release of lectins

A dual pH and glucose responsive boronic acid containing nanofiber was constructed for the reversible capture and release of lectins. The effects of surface groups and pH values on selective lectin capture were investigated by fluorescence microscopy. Compared to the pristine nanofibrous membrane, glucose and galactose functionalized nanofiber surfaces showed significantly higher capture of ConA and Jacalin, under alkaline conditions. On the other hand, treatment of the modified nanofibers with an acidic solution resulted in the detachment of both the lectins and glycopolymers from the nanofiber surface. As expected, once the glycopolymers are displaced, no lectins were adhered to the nanofiber surface under alkaline conditions. These functional nanofibers can therefore be easily modified and hence can be used for quick removal of selective proteins or toxins from the solution.

Two-component self-assembly of a tetra-guanidiniocarbonyl pyrrole cation and Na4EDTA: formation of pH switchable supramolecular networks.

A guanidiniocarbonyl pyrrole (GCP) cation forms stable H-bond assisted ion pairs with carboxylates even in aqueous solutions. A tetra GCP cation 1 undergoes efficient two-component self-assembly with Na4EDTA, a tetra-carboxylate, leading to 3D supramolecular networks. These networks show dual pH responsiveness and reversibly dissociate back into monomers upon addition of either acid or base.

Multiresponsive Microcapsules Based on Multilayer Assembly of Star Polyelectrolytes

Star polyelectrolytes (poly(N,N-dimethylaminoethyl methacrylate) (PDMAEMA)) with dual (temperature and pH) responsive properties were utilized to fabricate multiresponsive microcapsules via layer-by-layer (LbL) assembly. The LbL microcapsules are very robust and uniform, with higher stability and different internal structure compared with conventional microcapsules based on linear polyelectrolytes. Ionic strength in the polyelectrolyte solution during the microcapsule assembly process has a significant influence on the thickness and permeability of microcapsules. With increasing pH, the permeability of microcapsules decreases, and the transition from “open” to “closed” state for target molecules can be achieved within a narrow pH range (from pH 7 to 8). On the other hand, the overall size and permeability of the microcapsules decrease with increasing temperature (with a shrinkage of 54% in diameter at 60 °C compared with room temperature), thus allowing to reversibly load and unload the microcapsules with high efficiency. The organization and interaction of star polyelectrolytes within confined multilayer structure are the main driving forces for the responsiveness to external stimuli. The multiresponsive LbL microcapsules represent a novel category of smart microstructures as compared to traditional LbL microcapsules with “one-dimensional” response to a single stimulus, and they also have the potential to mimic the complex responsive microstructures found in nature and find applications in drug delivery, smart coatings, microreactors, and biosensors.

Dual photo- and pH-responsive supramolecular nanocarriers based on water-soluble pillar[6]arene and different azobenzene derivatives for intracellular anticancer drug delivery.

Two novel types of supramolecular nanocarriers fabricated by the amphiphilic host-guest inclusion complex formed from water-soluble pillar[6]arene (WP6) and azobenzene derivatives G1 or G2 have been developed, in which G1 is structurally similar to G2 but has an extra phenoxy group in its hydrophobic region. Supramolecular micelles can be initially formed by WP6 with G1, which gradually transform into layered structures with liquid-crystalline properties, whereas stable supramolecular vesicles are obtained from WP6 and G2, which exhibit dual photo- and pH-responsiveness. Notably, the resulting WP6⊃G2 vesicles can efficiently encapsulate anticancer drug mitoxantrone (MTZ) to achieve MTZ-loaded vesicles, which maintain good stability in a simulated normal physiological environment, whereas in an acid environment similar to that of tumor cells or with external UV irradiation, the encapsulated drug is promptly released. More importantly, cytotoxicity assay indicates that such vesicles have good biocompatibility and the MTZ-loaded vesicles exhibit comparable anticancer activity to free MTZ, especially with additional UV stimulus, whereas its cytotoxicity for normal cells was remarkably reduced. Flow cytometric analysis further confirms that the cancer cell death caused by MTZ-loaded vesicles is associated with apoptosis. Therefore, the dual pH- and UV-responsive supramolecular vesicles are a potential platform for controlled release and targeted anticancer drug delivery.

Dual pH‐ and temperature‐responsive hydrogels with extraordinary swelling/deswelling behavior and enhanced mechanical performances

ABSTRACTpH‐ and temperature‐responsive semi‐interpenetrating nanocomposite hydrogels (NC hydrogels) were prepared with surface‐functionalized graphene oxide (GO) as the crosslinker, N‐isopropylacrylamide (NIPAM) as the monomer, and chitosan (CS) as an additive. The effects of 3‐(trimethoxysilyl)propylmethacrylate‐modified GO sheets and CS content on various physical properties were investigated. Results show that PNIPAM/CS/GO hydrogels undergo a large volumetric change in response to temperature. Swelling ratios of PNIPAM/CS/GO hydrogels are much larger than those of the conventional organically crosslinked PNIPAM hydrogels. The deswelling test indicates that the deswelling rate was greatly enhanced by incorporating CS into the hydrogel network and using the surface‐functionalized GO as the crosslinker. The pH‐sensitivity of PNIPAM/CS/GO hydrogels is evident below their volume phase transition temperature. Moreover, the PNIPAM/CS/GO hydrogels have a much better mechanical property compared with traditional hydrogels even in a high water content of 90%. © 2014 Wiley Periodicals, Inc. J. Appl. Polym. Sci. 2015, 132, 41530.

Corner rounding of linear five-axis tool path by dual PH curves blending

The widespread linear five-axis tool path (G01 blocks) is usually described by two trajectories. One trajectory describes the position of the tool tip point, and the other one describes the position of the second point on the tool axis. The inherent disadvantages of linear tool path are tangential and curvature discontinuities at the corners in five-axis tool path, which will result in feedrate fluctuation and decrease due to the kinematic constraints of the machine tools. In this paper, by using a pair of quintic PH curves, a smoothing method is proposed to round the corners. There are two steps involved in our method. Firstly, according to the accuracy requirements of the tool tip contour and tool orientation tolerances, the corner is rounded with a pair of PH curves directly. Then, the control polygon lengths of PH curves are adjusted simply to guarantee the continuous variation of the tool orientation at the junctions between the transition curves and the remainder linear segments. Because the PH curves for corner rounding can be constructed without any iteration, and those two rounded trajectories are synchronized linearly in interpolation, which makes this smoothing method can be applied in a high efficiency way. Its high computational efficiency allows it to be implemented in real-time applications. This method has been integrated into a CNC system with an open architecture to implement on-line linear five-axis tool path smoothing. Simulations and experiments validate its practicability and reliability.

Proton-transfer reaction dynamics and energetics in calcification and decalcification.

CaCO3 -saturated saline waters at pH values below 8.5 are characterized by two stationary equilibrium states: reversible chemical calcification/decalcification associated with acid dissociation, Ca(2+) +HCO3 (-) ⇌CaCO3 +H(+) ; and reversible static physical precipitation/dissolution, Ca(2+) +CO3 (2-) ⇌CaCO3 . The former reversible reaction was determined using a strong base and acid titration. The saturation state described by the pH/PCO2 -independent solubility product, [Ca(2+) ][CO3 (2-) ], may not be observed at pH below 8.5 because [Ca(2+) ][CO3 (2-) ]/([Ca(2+) ][HCO3 (-) ]) ≪1. Since proton transfer dynamics controls all reversible acid dissociation reactions in saline waters, the concentrations of calcium ion and dissolved inorganic carbon (DIC) were expressed as a function of dual variables, pH and PCO2 . The negative impact of ocean acidification on marine calcifying organisms was confirmed by applying the experimental culture data of each PCO2 /pH-dependent coral polyp skeleton weight (Wskel) to the proton transfer idea. The skeleton formation of each coral polyp was performed in microspaces beneath its aboral ectoderm. This resulted in a decalcification of 14 weight %, a normalized CaCO3 saturation state Λ of 1.3 at PCO2 ≈400 ppm and pH ≈8.0, and serious decalcification of 45 % and Λ 2.5 at PCO2 ≈1000 ppm and pH ≈7.8.

Novel pH- and temperature-responsive polymer: Tertiary amine starch ether

A novel double pH- and temperature-responsive tertiary amine starch ether (TAS) has been developed. Synthesis was performed by grafting dipropyl or dibutyl epoxypropylamine onto hydroxyethyl starch. The cloud point temperatures (TC) of TAS could be tuned to a wide range from 26 to 72.8°C by changing the alkyl chain length, their average molar substitution (MS), and pH value of the solution. The TC of TAS increases with decreasing the alkyl chain length, MS, and pH value of the solution. A linear relationship occurs between the TC and the pH, indicating well-tunable TC. These TAS also showed single pH-sensitive property due to the existence of tertiary amino and hydrophobic alkyl groups. The synthetic strategy presented here could be employed in the preparation of other novel biomaterials with dual pH- and temperature-responsive properties from a variety of polysaccharides.

PH- and temperature-responsive hydrogels of acrylic acid, N-isopropylacrylamide and a non-ionic surfmer: phase behaviour, swelling properties and drug release

Copolymer hydrogels composed of N-isopropylacrylamide (NIPAM), acrylic acid (AA) and the non-ionic surfactant monomer (surfmer) ω-methoxy poly(ethylene oxide)40undecyl-α-methacrylate (PEO-R-MA-40) were prepared and studied with regard to swelling behaviour and drug release behaviour. The gels were prepared upon γ-irradiation of the corresponding aqueous comonomer solution in a one-step reaction. Transparent, stable hydrogels were obtained. Studies of light transmission indicate a dual pH- and T-responsive behaviour, which originates from the AA and NIPAM content of the gels, respectively. Presence of large amounts of surfmer increases the phase transition temperature, but also increases the network density, which lowers the permeability of the gels. Swelling properties and release of ibuprofen (Ibu) were studied in simulated gastric fluid (SGF, pH 1) and phosphate buffer solution (PBS, pH 6.8). It was found that swelling and release are controlled by the nature and quantity of comonomers, pH, temperature and ionic strength of the aqueous phase. Swollen gels shrink in SGF and PBS, whereas dry gels exhibit a strong swelling both in SGF and PBS. Copolymer gels of AA and surfmer exhibit a strong, linear release of Ibu in SGF and PBS. If NIPAM is copolymerized in the gel, the drug release is decelerated in SGF probably due to formation of hydrogen bonds between NIPAM and Ibu at low pH. For example, a gel composed of 10 % (w/w) NIPAM, 1 % (w/w) AA and 1.5 % (w/w) surfmer exhibits a release of 10 % within 2 h in SGF and 58 % within 20 h in PBS.

Preparation of pH- and salinity-responsive cellulose copolymer in ionic liquid

A novel biodegradable pH- and salinity-responsive cellulose copolymer was prepared by grafting 2-(Dimethylamino) ethylmethacrylate (DMAEMA) onto bagasse cellulose in ionic liquid. The grafting polymerization was achieved in 1-butyl-3-methylimidazolium chloride ([Bmim]Cl) under microwave irradiation. Copolymers were then characterized by Fourier transform infrared spectroscopy, scanning electron microscopy, X-ray diffraction and thermo gravimetric analysis measurements. The results revealed that polymer chains had been successfully bonded to the cellulose backbone. Furthermore, the self-assembly of cellulose-g-DMAEMA copolymers at various salt concentrations and pH solution were investigated by means of swelling behavior measurement. It indicated that the copolymers presented dual pH and salinity-responsive properties. The synthetic strategy showed great potential in the modification of other cellulosic biomass to afford new biomaterials with desired properties.

Dual thermo- and pH-sensitive poly(2-hydroxyethyl methacrylate-co-acrylic acid)-grafted graphene oxide

A two-step modification was used to attach atom transfer radical polymerization (ATRP) initiator onto graphene oxide surface. ATRP polymerization of 2-hydroxyethyl methacrylate (HEMA) was performed via “grafting from” approach. Due to uncontrolled ATRP of acrylic acid (AA), the Br-terminated P(HEMA) chains were converted to reversible addition–fragmentation chain transfer agent and polymerization of AA was done. The structure of modified nanosheets was characterized using X-ray diffraction analysis, Raman spectroscopy, proton nuclear magnetic resonance, scanning electron microscopy, and etc. These nanosheets showed dual pH- and thermo-sensitive properties as measured by UV–visible spectroscopy in different pH (2–13) and temperature (15–55 °C) values. Generally, UV absorbance of P(HEMA-co-AA)-grafted nanosheets was higher than P(HEMA)-grafted nanosheets. Also, it seems that the poly(acrylic acid) block induces more pH sensitivity behavior than P(HEMA) block. Lower critical solution temperature of polymer-grafted nanosheets were shifted to higher temperature when chain extension was performed.

Dual pH and temperature stimuli-responsive magnetic nanohydrogels for thermo-chemotherapy.

Dual stimuli pH and temperature-responsive nanohydrogels based on poly(N-isopropylacrylamide)-chitosan have been synthesized. Fe3O4 magnetic nanoparticles (NPs) (-12 nm) have been incorporated into hydrogels to achieve temperature optimized magnetic nanohydrogel (MNHG) for magnetic hyperthermia with lower critical solution temperature, LCST > 42 degrees C. The composite was further investigated for its potential application in drug delivery and in vitro cancer cell cytotoxicity. Water-bath assisted drug release studies were carried out using anti-cancer drug doxorubicin (DOX) in acetate buffer medium (pH - 4.6) to mimic tumor cell environment which is slightly acidic in nature. The pH and temperature responsiveness of the system was demonstrated by DOX release under different conditions. The released amount of DOX was found to be nearly 4 microg/mg above hyperthermia temperature (-42 degrees C) as opposed to only 1.9 microg/mg of MNHG at physiological temperature (37 degrees C) under acidic environment (pH - 4.6). Further, AC magnetic field (AMF) induced heating of NPs entrapped inside hydrogels showed appreciable reduction of cell population in human breast (MCF-7) and cervical carcinoma (HeLa) cell lines for given duration of field exposures. Quantitatively, death percentages of HeLa cells were nearly 35 and 45% while for MCF-7, these were 20 and 70% when exposed to AMF for 10 and 30 min, respectively. Further the cell killing efficacy of MNHG loaded with DOX was assessed under AMF using HeLa cell lines. The AMF induced heat triggered DOX release from the MNHG which enhances the cell death up to 85% due to combined effect of thermo-chemotherapeutics. The present system with both pH and temperature responsivity serves as a promising candidate for a combination therapy.

Synthesis of dual thermoresponsive and pH-sensitive hollow nanospheres by atom transfer radical polymerization

A facile and effective approach was developed to fabricate dual temperature- and pH-sensitive hollow nanospheres utilizing an atom transfer radical polymerization (ATRP) method. To do this silica nanoparticles were used as primary cores that could be etched by an hydrofluoric (HF) aqueous solution. Due to uncontrolled ATRP of acrylic acid (AA) methyl acrylate (MA) was polymerized via surface-initiated ATRP (SI-ATRP) a and poly(2-hydroxyethyl methacrylate) (PHEMA) block was added via the same approach. To synthesize poly(AA-co-HEMA)-grafted silica nanoparticles polymethyl acrylate (PMA) chains were hydrolyzed to polyacrylic acid (PAA) using an aqueous NaOH solution. PAA segments were partially crosslinked via an esterification reaction of –COOH groups with 1,4-butanediol. Finally, poly(AA-co-HEMA) hollow nanospheres were fabricated by etching silica cores with an HF aqueous solution. The structure of the nanospheres was revealed by transmission electron microscopy (TEM). These hollow nanospheres consisting of poly(AA-co-HEMA) in their structure showed dual pH- and thermo-sensitive properties as measured by dynamic light scattering (DLS). The hydrodynamic diameter was measured as an affected parameter under different pH (3–12) and temperature (25–55 °C) conditions. Results showed that by decreasing pH or by increasing temperature the hydrodynamic diameter decreased and a lower critical solution temperature (LCST) point was observed.

950 Dual Ambulatory pH Monitoring in Patients With Gastroesophageal Reflux Rendered Asymptomatic With Proton Pump Inhibitor Therapy

Background Studies have suggested that proton pump inhibitor (PPI) therapy in gastroesophageal reflux disease (GERD) achieves high rates of esophageal acid normalization.

Laryngopharyngeal reflux: paradigms for evaluation, diagnosis, and treatment.

This study aimed to describe current patterns for diagnosis and treatment of laryngopharyngeal reflux (LPR) and analyze differences between laryngologists and non-laryngologists. American Academy of Otolaryngology-Head and Neck Surgery and American Broncho-Esophagological Association members were invited to complete an online survey regarding evaluation, diagnosis, and treatment of LPR. Subgroup analysis was performed to identify differences between respondents who completed laryngology fellowships (LF) and those who did not (NL). Of 159 respondents, 40 were LF. Video documentation of laryngopharyngeal exams was almost universal among LF (97% vs 38%, P < .0001). Use of rigid (100%, P = .002) and flexible distal-chip technologies (94%, P = .004) was more common among LF. Diagnostic criteria were similar between the groups, with symptoms of heartburn, globus, and throat clearing thought most suggestive of LPR. Adjunctive tests most commonly used were barium esophagram and dual-probe pH testing with impedance. Laryngology fellowship-trained respondents used dual pH probes with impedance more often (P = .004). They were more likely to prescribe twice daily proton pump inhibitors with concurrent H2-blocker medication initially (P = .004) and to treat for longer than 4 weeks (P = .0003). Otolaryngologists are in agreement on symptoms and physical features of LPR; however, significant differences exist between laryngologists and non-laryngologists on the use of adjunctive testing and treatment strategies.

Chitosan/agarose hydrogels: Cooperative properties and microfluidic preparation

The preparation of composite biopolymer hydrogels offers the capability to produce biocompatible and biodegradable materials with cooperative properties. In this paper, two natural polymers, namely, chitosan and agarose were employed to prepare composite hydrogels with dual pH and temperature properties. The elastic modulus of the composite hydrogels increased with agarose concentration reaching the value of 1kPa for the chitosan/agarose gel with a 2% (w/v) concentration of agarose. In addition, composite gels exhibited a higher stability in acidic aqueous solutions, in comparison with agarose gels. The drug release properties of the composite hydrogels were tested by loading a model anticancer drug, 5-Fluorouracil, in the hydrogel interior. At pH=7.4, the cumulative release of 5-FU was ∼50% within 96h and decreased to ∼33% at pH=5.2, which was attributed to the different solubility of 5-FU as a function of pH. The preparation of composite microgels with controllable dimensions in the range from 42 to 18μm and with narrow size distribution (polidispersity not exceeding 1.5%) was achieved by the microfluidic emulsification of an aqueous mixture of chitosan and agarose and subsequent gelation of the precursor droplets by cooling.