Velocity-selective arterial spin labeling perfusion measurements in 2nd trimester human placenta with varying BMI.

INTRODUCTION: Proper placental development is crucial to fetal health but is challenging to functionally assess non-invasively and is thus poorly characterized in populations. Body mass index (BMI) has been linked with adverse outcomes, but the causative mechanism is uncertain. Velocity-selective arterial spin labeling (VS-ASL) MRI provides a method to non-invasively measure placental perfusion with robustness to confounding transit time delays. In this study, we report on the measurement of perfusion in the human placenta in early pregnancy using velocity-selective arterial spin labeling (VS-ASL) MRI, comparing non-obese and obese participants. METHODS: Participants (N = 97) undergoing routine prenatal care were recruited and imaged with structural and VS-ASL perfusion MRI at 15 and 21 weeks gestation. Resulting perfusion images were analyzed with respect to obesity based on BMI, gestational age, and the presence of adverse outcomes. RESULTS: At 15 weeks gestation BMI was not associated with placental perfusion or perfusion heterogeneity. However, at 21 weeks gestation BMI was associated with higher placental perfusion (p < 0.01) and a decrease in perfusion heterogeneity (p < 0.05). In alignment with past studies, perfusion values were also higher at 21 weeks compared to 15 weeks gestation. In a small cohort of participants with adverse outcomes, at 21 weeks lower perfusion was observed compared to participants with uncomplicated pregnancies. DISCUSSION: These results suggest low placental perfusion in the early second trimester may not be the culpable factor driving associations of obesity with adverse outcomes.

Lipid-based eye drop formulations for the management of evaporative dry eyes.

Dry eye disease is a progressive prevalent ocular surface disorder that arises from various factors and is characterized by insufficient quality and/or quantity of tears. The underlying pathophysiology is intricate and can progress to chronic, difficult-to-treat conditions. Multiple strategies and therapeutic approaches are utilized in its management that target one or more etiopathological components of dry eyes, which may include aqueous tear deficiency or evaporative dry eyes. The primary focus of this paper is on treatment alternatives that utilize lipids for the treatment of evaporative dry eyes. This may arise from either abnormal lipid production or inadequate lipid spreading caused by meibomian gland dysfunction. The hypothesis behind the development of these lipid-containing eye drops is that if they can imitate the lipid layer, they may be able to help in the management of the signs and symptoms of evaporative dry eyes. The lipids used in commercial formulations for dry eyes are mineral oil, castor oil, phospholipids, omega-3 fatty acid, and medium-chain triglycerides. The literature suggests the potential of lipid-containing eye drops to alleviate some of the signs and symptoms and enhance the quality of life for individuals suffering from evaporative dry eyes.

Chitosan nanoparticles laden contact lenses for enzyme-triggered controlled delivery of timolol maleate: A promising strategy for managing glaucoma.

To improve drug bioavailability, eye drops can be replaced by drug-eluting contact lenses. However, issues of drug leaching from lenses during manufacture and storage, and sterilization, currently limit their commercial application. To address the issues, stimuli-(lysozyme)-sensitive chitosan nanoparticles were developed to provide controlled ocular drug delivery. Nanoparticles were prepared by ionic gelation and characterized by TEM, X-ray diffraction, DSC, and FTIR. In the flux study, conventional-soaked contact lenses (SM-TM-CL) showed high-burst release, while with direct drug-only laden contact lenses (DL-TM-CL) the drug was lost during extraction and sterilization, as well as having poor swelling and optical properties. The nanoparticle-laden contact lenses (TM-Cht-NPs) showed controlled release of timolol for 120 h in the presence of lysozyme, with acceptable opto-physical properties. In the shelf-life study, the TM-Cht-NPs contact lenses showed no leaching or alteration in the drug release pattern. In animal studies, the TM-NPs-CL lenses gave a high drug concentration in rabbit tear fluid (mean = 11.01 µg/mL for 56 h) and helped maintain a low intraocular pressure for 120 h. In conclusion, the chitosan nanoparticle-laden contact lenses demonstrated the potential application to treat glaucoma with acceptable opto-physical properties and addressed the issues of drug-leaching during sterilization and storage.

Cotyledon-Specific Flow Evaluation of Rhesus Macaque Placental Injury Using Ferumoxytol Dynamic Contrast-Enhanced MRI.

BACKGROUND: Recently, dynamic contrast-enhanced (DCE) MRI with ferumoxytol as contrast agent has recently been introduced for the noninvasive assessment of placental structure and function throughout. However, it has not been demonstrated under pathological conditions. PURPOSE: To measure cotyledon-specific rhesus macaque maternal placental blood flow using ferumoxytol DCE MRI in a novel animal model for local placental injury. STUDY TYPE: Prospective animal model. SUBJECTS: Placental injections of Tisseel (three with 0.5 mL and two with 1.5 mL), monocyte chemoattractant protein 1 (three with 100 µg), and three with saline as controls were performed in a total of 11 rhesus macaque pregnancies at approximate gestational day (GD 101). DCE MRI scans were performed prior (GD 100) and after (GD 115 and GD 145) the injection (term = GD 165). FIELD STRENGTH/SEQUENCE: 3 T, T1-weighted spoiled gradient echo sequence (product sequence, DISCO). ASSESSMENT: Source images were inspected for motion artefacts from the mother or fetus. Placenta segmentation and DCE processing were performed for the dynamic image series to measure cotyledon specific volume, flow, and normalized flow. Overall placental histopathology was conducted for controls, Tisseel, and MCP-1 animals and regions of tissue infarctions and necrosis were documented. Visual inspections for potential necrotic tissue were conducted for the two Tisseelx3 animals. STATISTICAL TESTS: Wilcoxon rank sum test, significance level P < 0.05. RESULTS: No motion artefacts were observed. For the group treated with 1.5 mL of Tisseel, significantly lower cotyledon volume, flow, and normalized flow per cotyledon were observed for the third gestational time point of imaging (day ~145), with mean normalized flow of 0.53 minute-1 . Preliminary histopathological analysis shows areas of tissue necrosis from a selected cotyledon in one Tisseel-treated (single dose) animal and both Tisseelx3 (triple dose) animals. DATA CONCLUSION: This study demonstrates the feasibility of cotyledon-specific functional analysis at multiple gestational time points and injury detection in a placental rhesus macaque model through ferumoxytol-enhanced DCE MRI. LEVEL OF EVIDENCE: NA TECHNICAL EFFICACY: Stage 2.

The C5aR1 complement receptor: A novel immunomodulator of insulin action in skeletal muscle.

The complement system constitutes an integral component of the innate immune system and plays a critical role in adaptive immunity. Activation of this system engenders the production of complement peptide fragments, including C5a, which engage G-protein coupled receptors predominantly expressed in immune-associated cells, such as neutrophils, initiating pro-inflammatory responses. Intriguingly, our investigation has unveiled the presence of C5a receptor 1 (C5aR1) expression within skeletal muscle, a key metabolic tissue and primary target of insulin. Herein, we demonstrate that C5aR1 activation by C5a in differentiated human skeletal muscle cells elicits acute suppression of insulin signalling. This suppression manifests as impaired insulin-dependent association between IRS1 and the p85 subunit of PI3-kinase, a 50% reduction in Akt phosphorylation, and a 60% decline in insulin-stimulated glucose uptake. This impairment in insulin signalling is associated with a three-fold elevation in intramyocellular diacylglycerol (DAG) levels and a two-fold increase in cytosolic calcium content, which promote PKC-mediated IRS1 inhibition via enhanced phosphorylation at IRS1 Ser1101. Significantly, our findings demonstrate that structurally diverse C5aR1 antagonists, along with genetic deletion or stable silencing of C5aR1 by 80% using short-hairpin RNA, effectively attenuate repression of insulin signalling by C5a in LHCN-M2 human skeletal myotubes. These results underscore the potential of heightened C5aR1 activation, characteristic of obesity and chronic inflammatory conditions, to detrimentally impact insulin function within skeletal muscle cells. Additionally, the study suggests that agents targeting the C5a-C5aR axis, originally devised for mitigating complement-dependent inflammatory conditions, may offer therapeutic avenues to ameliorate immune-driven insulin resistance in key peripheral metabolic tissues, including skeletal muscle.

Src kinase partially mediates cytokine-induced endothelial dysfunction.

OBJECTIVES: Endothelial dysfunction is known to be a key characteristic of preeclampsia (PE) and can contribute to progression of symptoms and injury to multiple organ systems. Delivery is the only treatment for progression of PE, but development of an endothelial-based therapy for PE presents a promising strategy. Growth factors and cytokines are dysregulated in PE and can impact endothelial function, manifesting changes in Ca2+ signaling and interruptions in monolayer barrier function that contribute to symptoms of hypertension, proteinuria, and edema. In this study, we highlight Src kinase as a partial mediator of growth factor and cytokine mediated endothelial dysfunction. STUDY DESIGN: Fura-2 Ca2+ imaging and Electrical Cell Impedance Sensing (ECIS) assays are performed on growth factor or cytokine exposed human umbilical vein endothelial cells (HUVECs). Inhibitors to MEK/ERK (U0126) or Src (PP2) are used to determine the contribution of kinase signaling pathways. MAIN OUTCOME MEASURES: Decreases in HUVEC Ca2+ signaling or monolayer resistance measure endothelial dysfunction. Reversal of endothelial dysfunction by kinase inhibitors reveals the respective contibutions of MEK/ERK and Src kinase. RESULTS: We show that Src inhibition protects Ca2+ signaling responses against insults induced by VEGF165, bFGF, PlGF, TNFα, and IL-1ß. Additionally, we show that Src inhibition protects the endothelial monolayer from the full impact of TNFα insult. Further, we find that MEK/ERK inhibition does not offer protection from growth factor-mediated endothelial dysfunction. CONCLUSIONS: The results of this study suggest cytokine and growth factor-stimulated Src kinase plays a partial role on promoting endothelial dysfunction in HUVECs.

Caveolin-3 loss linked with the P104L LGMD-1C mutation modulates skeletal muscle mTORC1 signalling and cholesterol homeostasis.

BACKGROUND: Caveolins are the principal structural components of plasma membrane caveolae. Dominant pathogenic mutations in the muscle-specific caveolin-3 (Cav3) gene isoform, such as the limb girdle muscular dystrophy type 1C (LGMD-1C) P104L mutation, result in dramatic loss of the Cav3 protein and pathophysiological muscle weakness/wasting. We hypothesize that such muscle degeneration may be linked to disturbances in signalling events that impact protein turnover. Herein, we report studies assessing the effects of Cav3 deficiency on mammalian or mechanistic target of rapamycin complex 1 (mTORC1) signalling in skeletal muscle cells. METHODS: L6 myoblasts were stably transfected with Cav3P104L or expression of native Cav3 was abolished by CRISPR/Cas9 genome editing (Cav3 knockout [Cav3KO]) prior to performing subcellular fractionation and immunoblotting, analysis of real-time mitochondrial respiration or fixed cell immunocytochemistry. Skeletal muscle from wild-type and Cav3-/- mice was processed for immunoblot analysis of downstream mTORC1 substrate phosphorylation. RESULTS: Cav3 was detected in lysosomal-enriched membranes isolated from L6 myoblasts and observed by confocal microscopy to co-localize with lysosomal-specific markers. Cav3P104L expression, which results in significant (~95%) loss of native Cav3, or CRISPR/Cas9-mediated Cav3KO, reduced amino acid-dependent mTORC1 activation. The decline in mTORC1-directed signalling was detected by immunoblot analysis of L6 muscle cells and gastrocnemius Cav3-/- mouse muscle as judged by reduced phosphorylation of mTORC1 substrates that play key roles in the initiation of protein synthesis (4EBP1S65 and S6K1T389 ). S6K1T389 and 4EBP1S65 phosphorylation reduced by over 75% and 80% in Cav3KO muscle cells and by over 90% and 30% in Cav3-/- mouse skeletal muscle, respectively. The reduction in protein synthetic capacity in L6 muscle cells was confirmed by analysis of puromycylated peptides using the SUnSET assay. Cav3 loss was also associated with a 26% increase in lysosomal cholesterol, and pharmacological manipulation of lysosomal cholesterol was effective in replicating the reduction in mTORC1 activity observed in Cav3KO cells. Notably, re-expression of Cav3 in Cav3KO myoblasts normalized lysosomal cholesterol content, which coincided with a recovery in protein translation and an associated increase in mTORC1-directed phosphorylation of downstream targets. CONCLUSIONS: Our findings indicate that Cav3 can localize on lysosomal membranes and is a novel regulator of mTORC1 signalling in muscle. Cav3 deficiency associated with the Cav3P104L mutation impairs mTORC1 activation and protein synthetic capacity in skeletal muscle cells, which may be linked to disturbances in lysosomal cholesterol trafficking and contribute to the pathology of LGMD-1C.

Efficacy and safety of zinc in the prevention of oral mucositis in children with cancer receiving intensified chemotherapy: A randomized double-blind placebo-controlled trial.

BACKGROUND AND AIMS: A limited number of safe and effective preventive options for oral mucositis (OM) are available. This randomized, double-blind, placebo-controlled trial aimed to evaluate the efficacy and safety of zinc in preventing OM in children with cancer receiving intensified chemotherapy. METHODS: Children aged 3-18 years were randomized to receive oral zinc at 1 mg/kg/dose daily for 14 days or a placebo at the same doses and schedule. The primary outcome of this study was to determine the effect of oral zinc in the prevention of OM, and secondary outcomes included any adverse effect of oral zinc, the severity and duration of OM, and the need for hospitalizations. RESULTS: A total of 90 children were randomized to either the oral zinc (n = 44) or placebo group (n = 46). The incidence of OM in the zinc group was 20.5%, while that in the placebo group was 19.6% (p = .91; risk ratio: 1.04, 95% CI 0.45-2.30). There were no significant adverse events of the drug observed. There were no significant differences between the two groups in the severity (p = .79), the mean time of onset (p = .09), the mean duration of OM (p = .18), and the need for hospitalizations (p = 1.0). CONCLUSIONS: Among children on cancer chemotherapy, there was no decrease in the incidence of OM observed with oral zinc at a dose of 1 mg/kg/day. No significant adverse events were observed with administering oral zinc. Further research is warranted to test higher doses of oral zinc with longer duration for a clinically significant effect.

Ferumoxytol dynamic contrast enhanced magnetic resonance imaging identifies altered placental cotyledon perfusion in rhesus macaques†.

Identification of placental dysfunction in early pregnancy with noninvasive imaging could be a valuable tool for assessing maternal and fetal risk. Dynamic contrast enhanced (DCE) magnetic resonance imaging (MRI) can be a powerful tool for interrogating placenta health. After inoculation with Zika virus or sham inoculation at gestation age (GA) 45 or 55 days, animals were imaged up to three times at GA65, GA100, and GA145. DCE MRI images were acquired at all imaging sessions using ferumoxytol, an iron nanoparticle-based contrast agent, and analyzed for placental intervillous blood flow, number of perfusion domains, and perfusion domain volume. Cesarean section was performed at GA155, and the placenta was photographed and dissected for histopathology. Photographs were used to align cotyledons with estimated perfusion domains from MRI, allowing comparison of estimated cotyledon volume to pathology. Monkeys were separated into high and low pathology groups based on the average number of pathologies present in the placenta. Perfusion domain flow, volume, and number increased through gestation, and total blood flow increased with gestation for both low pathology and high pathology groups. A statistically significant decrease in perfusion domain volume associated with pathology was detected at all gestational ages. Individual perfusion domain flow comparisons demonstrated a statistically significant decrease with pathology at GA100 and GA145, but not GA65. Since ferumoxytol is currently used to treat anemia during human pregnancy and as an off-label MRI contrast agent, future transition of this work to human pregnancy may be possible.

In vitro and in vivo evaluation of cyclosporine-graphene oxide laden hydrogel contact lenses.

Drug-eluting contact lens can substitute the multiple eye drop therapy. However, loading hydrophobic drug like cyclosporine in the contact lens is very challenging, due to low drug uptake (via soaking method); and alteration in the swelling and optical properties which restricts its clinical application. To address the above issues, graphene oxide (GO, large surface area with oxygen containing functional groups) was incorporated in the contact lenses during fabrication. These GO-laden contact lenses (SM-GO-Cys) as well as blank contact lenses (SM-Cys) were soaked in the solution of cyclosporine. Alternatively, cyclosporine-laden contact lenses (DL-Cys-20) and cyclosporine-GO-laden contact lenses (DL-Cys-20-GO) were fabricated by adding drug and drug-GO (at various level of GO) during fabrication, respectively. Contact angle and swelling data showed increase in water holding capacity of GO laden contact lenses. Optical property was significantly improved due to molecular dispersion of drug on the surface of GO sheets. The drug uptake and in vitro release profile was improved with GO-laden contact lenses by soaking method (SM-GO-Cys-400n) due to hydrophobic interactions between GO and drug. Adding cyclosporine-GO (DL-Cys-20-GO-800n) during fabrication significantly improved drug release kinetics with higher drug leaching (during extraction and sterilization) due to increased swelling, improved dissolution and molecular dispersion of drug on GO sheets. Ocular irritation and histopathological studies demonstrated the safety of GO-contact lens. The in vivo drug release studies in the rabbit eye showed significant improvement in mean residence time (MRT) and area under the curve (AUC) using DL-Cys-20-GO-800n contact lens compared to eye drop solution with reduction in protein adherence value. The study demonstrated that the incorporation of GO into the contact lens can control the release of cyclosporine as well as improved the lens swelling and transmittance properties.

Application of fungal copper carbonate nanoparticles as environmental catalysts: organic dye degradation and chromate removal.

Biomineralization is a ubiquitous process in organisms to produce biominerals, and a wide range of metallic nanoscale minerals can be produced as a consequence of the interactions of micro-organisms with metals and minerals. Copper-bearing nanoparticles produced by biomineralization mechanisms have a variety of applications due to their remarkable catalytic efficiency, antibacterial properties and low production cost. In this study, we demonstrate the biotechnological potential of copper carbonate nanoparticles (CuNPs) synthesized using a carbonate-enriched biomass-free ureolytic fungal spent culture supernatant. The efficiency of the CuNPs in pollutant remediation was investigated using a dye (methyl red) and a toxic metal oxyanion, chromate Cr(VI). The biogenic CuNPs exhibited excellent catalytic properties in a Fenton-like reaction to degrade methyl red, and efficiently removed Cr(VI) from solution due to both adsorption and reduction of Cr(VI). X-ray photoelectron spectroscopy (XPS) identified the oxidation of reducing Cu species of the CuNPs during the reaction with Cr(VI). This work shows that urease-positive fungi can play an important role not only in the biorecovery of metals through the production of insoluble nanoscale carbonates, but also provides novel and simple strategies for the preparation of sustainable nanomineral products with catalytic properties applicable to the bioremediation of organic and metallic pollutants, solely and in mixtures.

Advances and challenges in the nanoparticles-laden contact lenses for ocular drug delivery.

The delivery of drugs that target ocular tissues is challenging due to the physiological barriers of the eye like tear dilution, nasolacrimal drainage, blinking, tear turnover rate and low residence time Drug-laden contact lenses can be a possible solution to overcome some of these challenges. Nanoparticles are being extensively studied as novel systems for loading drugs into therapeutic contact lenses. The versatile features of the organic and inorganic nanoparticles and their diverse physicochemical properties make it possible to load and sustain drug release from the contact lenses. Nevertheless, several issues remains to be solved before its clinical application and commercialization such as changes in contact lens swelling (water content), transmittance, protein adherence, surface roughness, tensile strength, ion and oxygen permeability and drug leaching during contact lens manufacture. However, clinical studies demonstrated the potential of therapeutic contact lenses to manage the scientific, commercial and regulatory challenges to make its place in the market. This review highlights the different methodologies used to fabricate nanoparticle-laden contact lenses and highlights the major advances and challenges to commercialization.

Controlled bimatoprost release from graphene oxide laden contact lenses: In vitro and in vivo studies.

Ocular drug delivery using contact lenses may be able to substitute for eye drop therapy. However, issues with hydrophobic drugs (like bimatoprost that is used to treat glaucoma) such as low drug uptake using a simple soaking method into preformed contact lenses and alteration in the swelling and transmittance of lenses restricts the application for drug delivery. This research uses graphene oxide (GO) to control the release of bimatoprost from contact lenses along with improvements in the drug uptake, and lens swelling and transmittance. GO was loaded into silicone hydrogel contact lenses by adding the GO at the same time as lenses were polymerized. These lenses were soaked in bimatoprost. Alternatively contact lenses, either with or without GO, were produced by adding bimatoprost during lens polymerization. GO improved contact lens swelling due to its water binding capacity and lens transmittance due to the molecular dispersion of bimatoprost on the surface of the GO which prevented the local precipitation of the drug. The bimatoprost uptake was not improved in the presence of GO. However, its in vitro release profile was improved. Adding bimatoprost and GO at the same time as lenses were polymerized (DL-GO-BMT) significantly decreased the loss of drug during extraction and sterilization in comparison to contact lenses (DL-BMT) without GO. As the amount of GO was increased, the DL-GO-BMT lenses showed a significant decrease in the burst and cumulative release of bimatoprost. Ocular irritation and histopathology reports demonstrated the safety of GO contact lens. The in vivo pharmacokinetic studies in the rabbit tear fluid showed significant improvement in mean residence time (MRT) and area under the curve (AUC) with DL-GO-0.2 µg-BMT-100 contact lens in comparison to eye drop solution. The study demonstrated that the addition of GO to contact lenses can control the release of bimatoprost as well as improved the lens swelling and transmittance. However, further optimization is needed to modulate the release of drug within the therapeutic level to manage glaucoma.

Recent advances in ophthalmic preparations: Ocular barriers, dosage forms and routes of administration.

The human eye is a complex organ with unique anatomy and physiology that restricts the delivery of drugs to target ocular tissues/sites. Recent advances in the field of pharmacy, biotechnology and material science have led to development of novel ophthalmic dosage forms which can provide sustained drug delivery, reduce dosing frequency and improve the ocular bioavailability of drugs. This review highlights the different anatomical and physiological factors which affect ocular bioavailability of drugs and explores advancements from 2016 to 2020 in various ophthalmic preparations. Different routes of drug administration such as topical, intravitreal, intraocular, juxtascleral, subconjunctival, intracameral and retrobulbar are discussed with their advances and limitations.

Thermoresponsive liquid crystalline formulation of Exemestane: Design and structural characterization.

Exemestane (EXE), a drug used for the treatment of breast cancer, has limited aqueous solubility of 0.08 mg/mL and log P∼ 4.22. The only available marketed formulation in form of tablets possess limitations of poor oral absorption (∼ 42 %), low solubility, extensive hepatic metabolism and numerous adverse effects due to its peripheral absorption. In order to address these issues, an alternative route of topical application is attempted through a lamellar liquid crystal based formulation. Pluronic® was used as stabilizer due to its higher surface activity and gelling properties. The solubility enhancement of EXE was achieved using liquid crystal formulation. We have investigated the effect of concentration of oil, Smix (surfactant - cosurfactant mixture) and EXE on lattice parameter, rheology and drug release for various combinations of the formulation. The small angle x-ray scattering (SAXS) measurement demonstrated an evidence of a lamellar structure with lattice parameter ∼15 nm, which increases with corresponding increase in oil and EXE due to increase in hydrophobic interactions leading to an expansion of lamella. The inter lamellar distance decreases at higher surfactant concentration, due to the distribution of the same amount of oil and drug within larger concentration of surfactant molecules. The rheology measurement exhibited gel like properties at low shear rate indicating soft gel formation, which converts to Newtonian type flowing liquid at higher shear rate. At constant Smix with increasing oil content, the viscosity decreases, which is attributed to the dilution of the lamellar structures with oil. The temperature sweep rheology reveals a change in the viscosity near physiological temperature, which may be attributed to the structural transition of lamellae. The formulation remains gel like at room temperature, which aids in proper application to skin and converts it to free flowing liquid above 37 °C. The invitro drug release of optimized formulation for 24 h was ∼ 38 % at 37 °C, which increased to 50 % at 42 °C. Accordingly, this formulation containing thermoresponsive lamellar liquid crystal gels of EXE represents a viable option for hyperthermia induced enhanced drug release. The characteristic and advantageous features offered by this formulation includes improved bioavailability of EXE due to enhanced solubility, permeability and absorption.

Tailored Doxycycline Hyclate Loaded In Situ Gel for the Treatment of Periodontitis: Optimization, In Vitro Characterization, and Antimicrobial Studies.

Currently, periodontitis is treated by oral dosage forms (antibiotics) which shows systemic side effects and failed to reach the therapeutic concentration (above minimum inhibitory concentration, MIC) in the periodontal pocket. The present study aimed to overcome the above issues, by designing tailored doxycycline hyclate laden in situ gel by Poloxamer 407, chitosan, and polyethylene glycol 600. The in situ gel-forming system has attracted attention owing to its ability of sustained drug release above MIC, easy administration (syringeability), and high drug retention (localization) in the periodontal cavity. The Box-Behnken design (BBD) was used to tailor and optimize the concentration of Poloxamer 407 (X1 = 14.3%), chitosan (X2 = 0.58%), and polyethylene glycol 600 (X3 = 1.14%) to achieve sufficient syringeability (149 N), t90% (1105 min), and viscosity at non-physiological condition (512 cps) and physiological condition (5415 cps). The optimized in situ gel was clear and isotonic (RBCs test). The gelation temperature of the optimized in situ was 34 ± 1°C with sufficient mucoadhesive strength (26 ± 2 dyn/cm2), gel strength (29 ± 2 sec), and texture profile for periodontal application. The in vitro drug release studies showed sustain release from optimized in situ gel (24h) in comparison to marketed gel (7h). The antimicrobial activity (cup plate technique) of the in situ gel was equivalent to the marketed doxycycline gel, which suggests that the doxycycline hyclate retained its antimicrobial efficacy when formulated as in situ gelling system. In conclusion, BBD was effectively utilized to optimize in situ gel with minimum level of polymers to achieve the required characteristics of the in situ gel for sustaining drug delivery to treat periodontitis.

Oral bioavailability improvement of felodipine using tailored microemulsion: Surface science, ex vivo and in vivo studies.

Felodipine is a calcium channel blocker, which shows low oral bioavailability (<15%) owing to poor water solubility and high first pass metabolism. The aim of the present investigation was to study the surface science (dynamic surface tension) and characteristics of microemulsion (Capmul MCM, Tween 20 and polyethylene glycol) to enhance the oral bioavailability of felodipine by improving permeability of the drug in the intestine. The paper is the first attempt to study the stability of oil-water interface of microemulsion using bubble tensiometer. The Smix at 2:1 ratio showed the maximum microemulsion area which did not alter in the presence of drug. The microemulsion batch coded Fe-O5-Smix45 (5% Capmul MCM and 45% Smix) was selected based on transmittance (>99%), dilution (stable after 100 times dilution with water), size (15.1 nm), dispersibility (grade A) and thermodynamic stability studies. The dynamic surface tension at newly created surface indicate the stability of surfactant film at the oil/water interface. The microemulsion was also stable in the presence of drug and in different buffer phases. The ex vivo intestinal permeability studies showed significant increase in the microemulsion permeation (74.1% after 1 h) in comparison to the felodipine suspension (16.9% after 1 h). The in vivo pharmacokinetic parameters in the rat model confirmed the improvement in oral bioavailability with microemulsion (relative bioavailability = 21.9) in comparison to the felodipine suspension, due to high surface area of oil droplets and its lymphatic uptake via transcellular route. In conclusion, the stable microemulsion offers a promising approach to improve the oral bioavailability of felodipine which can help to reduce the dose and its associated side effects.

Cytokine concentrations direct endothelial function in pregnancy and preeclampsia.

Endothelial dysfunction is a prominent feature of preeclampsia, a hypertensive disorder of pregnancy, and contributes to multiple symptoms characteristic of the syndrome. A myriad of growth factors and cytokines are dysregulated in preeclampsia as compared to normal pregnancy, however, a complete appreciation of the effect of changing concentrations of these factors on endothelial function is lacking. In this study, we evaluate the effect of a variety of growth factors and cytokines on Ca2+ signaling and monolayer integrity. We report that VEGF165, TNFα, EGF, and IL-1ß either improve or inhibit Ca2+ signaling depending on dose, whereas TNFα and IL-1ß reduce monolayer integrity and bFGF increases monolayer integrity. Additionally, to model the effects of combinations of growth factors and cytokines, we screened for Ca2+ signaling changes in response to 16 dose combinations of VEGF165 and TNFα together. This revealed an optimal combination capable of supporting pregnancy-adapted Ca2+ signaling, and that changes in either VEGF165 or TNFα dose would result in a shift toward suppressed function. This study shows in detail how growth factor or cytokine concentration effects endothelial cell function. Such data can be used to model how changing growth factor and cytokine levels in normal pregnancy may contribute to healthy endothelial function and in preeclampsia may promote endothelial dysfunction. The results of VEGF165 and TNFα combination treatments suggest that more complex growth factor and cytokine combination modeling may be important in order to more accurately understand the effects of circulating factors on the endothelial function.

Role of Protein in Fungal Biomineralization of Copper Carbonate Nanoparticles.

Biomineralization processes are of key importance in the biogeochemical cycling of metals and other elements by microorganisms, and several studies have highlighted the potential applications of nanoparticle synthesis via biomineralization. The roles played by proteins in the transformation and biologically induced biomineralization of metals by microorganisms is not well understood, despite the interactions of protein and nanoparticles at mineral interfaces attracting much interest in various emerging fields for novel biomaterial synthesis. Here, we have elucidated the association and involvement of fungal proteins in the formation of biogenic copper carbonate nanoparticles (CuNPs) using a carbonate-enriched biomass-free ureolytic fungal culture supernatant. Proteomic analysis was conducted that identified the major proteins present in the culture supernatant. Of the proteins identified, triosephosphate isomerase (TPI) exhibited a strong affinity to the CuNPs, and the impact of purified TPI on CuNP formation was studied in detail. The combined use of scanning electron microscopy (SEM) and confocal laser scanning microscopy (CLSM) confirmed that TPI played an important role in controlling the morphology and structure of the nanomaterials. Fourier transform infrared spectroscopy (FTIR) was applied to examine conformational changes of the proteins to further clarity the interaction mechanisms with CuNPs during biomineralization. Such analyses revealed unfolding of proteins on the mineral surface and an increase in ß sheets within the protein structure. These results extend understanding of how microbial systems can influence biomineral formation through protein secretion, the mechanisms involved in formation of complex protein/inorganic systems, and provide useful guidelines for the synthesis of inorganic-protein based nanomaterials.