As a topical delivery system, a nanoscaled emulsion is considered a good carrier of several active ingredients that convey several side effects upon oral administration, such as nonsteroidal anti-inflammatory drugs (NSAIDs).
We investigated the in vitro permeation properties and the in vivo pharmacodynamic activities of different nanoscaled emulsions containing ibuprofen, an NSAID, as an active ingredient and newly synthesized palm olein esters (POEs) as the oil phase.
A ratio of 25:37:38 of oil phase:aqueous phase:surfactant was used, and different additives were used for the production of a range of nanoscaled emulsions. Carbopol® 940 dispersion neutralized by triethanolamine was employed as a rheology modifier. In some circumstances, menthol and limonene were employed at different concentrations as permeation promoters. All formulae were assessed in vitro using Franz diffusion cell fitted with full-thickness rat skin. This was followed by in vivo evaluation of the anti-inflammatory and analgesic activities of the promising formulae and comparison of the effects with that of the commercially available gel.
Results and discussion:
Among all other formulae, formula G40 (Carbopol® 940-free formula) had a superior ability in transferring ibuprofen topically compared with the reference. Carbopol® 940 significantly decreased the amount of drug transferred from formula G41 through the skin as a result of swelling, gel formation, and reduction in drug thermodynamic activity. Nonetheless, the addition of 10% w/w of menthol and limonene successfully overcame this drawback since, relative to the reference, higher amount of ibuprofen was transferred through the skin. By contrast, these results were relatively comparable to that of formula G40. Pharmacodynamically, the G40, G45, and G47 formulae exhibited the highest anti-inflammatory and analgesic effects compared with other formulae.
The ingredients and the physical properties of the nanoscaled emulsions produced by using the newly synthesized POEs succeeded to deliver ibuprofen competently.
in vivo analgesic; anti-inflammatory effects
Background and purpose:
Chemokines play a critical role in the pathogenesis of asthma and facilitate the recruitment of inflammatory cells in the airways. Evidence now suggests that airway smooth muscle (ASM) may serve as a source of chemokines in inflamed airways. Although vitamin D has potent anti-inflammatory properties in vitro in some cell types, its effects on ASM cells remain unclear. Here, we investigated whether 1α, 25-dihydroxy vitamin D3 (calcitriol) modulated chemokine production in ASM.
Human ASM cell cultures were derived from tracheal samples taken during surgery. ASM cells were treated with tumour necrosis factor alpha (TNFα) and/or interferon gamma (IFNγ) for 24 h in the presence of calcitriol and/or the glucocorticoid fluticasone added 2 h before. RANTES (regulated upon activation, normal T-cell expressed and secreted), interferon-inducible protein 10 (IP-10) and fractalkine (FKN) levels in cell supernatants were measured by ELISA.
In TNFα-treated cells, calcitriol inhibited RANTES and IP-10 secretion in a concentration-dependent manner. FKN levels were negligible. In TNFα/IFNγ-treated cells, whereas fluticasone or calcitriol alone partially inhibited RANTES secretion (by 38 and 20%, respectively), the combination of both drugs additively inhibited RANTES secretion (by 60%). No effect was observed on IP-10 secretion. Whereas fluticasone enhanced FKN secretion (by 50%), calcitriol significantly decreased FKN levels (by 50%). Interestingly, calcitriol blocked the stimulatory effect of fluticasone on FKN secretion, which was inhibited by 60% with the combination of calcitriol and fluticasone.
Conclusions and implications:
These findings suggest that vitamin D uniquely modulates human ASM expression of chemokines and may exert some beneficial effects in the treatment of steroid-resistant patients with asthma.
chemokines; cytokines; glucocorticoids; steroid resistance; asthma; airway structural cells; inflammation; gene expression; drug; calcitriol
Fibroproliferative airway remodelling, including increased airway smooth muscle (ASM) mass and contractility, contributes to airway hyperresponsiveness in asthma. In vitro studies have shown that maturation of ASM cells to a (hyper)contractile phenotype is dependent on laminin, which can be inhibited by the laminin-competing peptide Tyr-Ile-Gly-Ser-Arg (YIGSR). The role of laminins in ASM remodelling in chronic asthma in vivo, however, has not yet been established.
Using an established guinea pig model of allergic asthma, we investigated the effects of topical treatment of the airways with YIGSR on features of airway remodelling induced by repeated allergen challenge, including ASM hyperplasia and hypercontractility, inflammation and fibrosis. Human ASM cells were used to investigate the direct effects of YIGSR on ASM proliferation in vitro.
Topical administration of YIGSR attenuated allergen-induced ASM hyperplasia and pulmonary expression of the proliferative marker proliferating cell nuclear antigen (PCNA). Treatment with YIGSR also increased both the expression of sm-MHC and ASM contractility in saline- and allergen-challenged animals; this suggests that treatment with the laminin-competing peptide YIGSR mimics rather than inhibits laminin function in vivo. In addition, treatment with YIGSR increased allergen-induced fibrosis and submucosal eosinophilia. Immobilized YIGSR concentration-dependently reduced PDGF-induced proliferation of cultured ASM to a similar extent as laminin-coated culture plates. Notably, the effects of both immobilized YIGSR and laminin were antagonized by soluble YIGSR.
These results indicate that the laminin-competing peptide YIGSR promotes a contractile, hypoproliferative ASM phenotype in vivo, an effect that appears to be linked to the microenvironment in which the cells are exposed to the peptide.
Asthma is characterized by the association of airway hyperresponsiveness (AHR), inflammation, and remodelling. The aim of the present article is to review the pivotal role of airway smooth muscle (ASM) in the pathophysiology of asthma. ASM is the main effector of AHR. The mechanisms of AHR in asthma may involve a larger release of contractile mediators and/or a lower release of relaxant mediators, an improved ASM cell excitation/contraction coupling, and/or an alteration in the contraction/load coupling. Beyond its contractile function, ASM is also involved in bronchial inflammation and remodelling. Whereas ASM is a target of the inflammatory process, it can also display proinflammatory and immunomodulatory functions, through its synthetic properties and the expression of a wide range of cell surface molecules. ASM remodelling represents a key feature of asthmatic bronchial remodelling. ASM also plays a role in promoting complementary airway structural alterations, in particular by its synthetic function.
In ocular tissue, arachidonic acid is metabolized by cyclooxygenase to prostaglandins which are the most important lipid derived mediators of inflammation. Presently nonsteroidal anti-inflammatory drugs (NSAIDs) which are cyclooxygenase (COX) inhibitors are being used for the treatment of inflammatory disorders. NSAIDs used in ophthalmology, topically, are salicylic-, indole acetic-, aryl acetic-, aryl propionic- and enolic acid derivatives. NSAIDs are weak acids with pKa mostly between 3.5 and 4.5, and are poorly soluble in water. Aqueous ophthalmic solutions of NSAIDs have been made using sodium, potassium, tromethamine and lysine salts or complexing with cyclodextrins/solubilizer. Ocular penetration of NSAID demands an acidic ophthalmic solution where cyclodextrin could prevent precipitation of drug and minimize its ocular irritation potential. The incompatibility of NSAID with benzalkonium chloride is avoided by using polysorbate 80, cyclodextrins or tromethamine. Lysine salts and α-tocopheryl polyethylene glycol succinate disrupt corneal integrity, and their use requires caution. Thus a nonirritating ophthalmic solution of NSAID could be formulated by dissolving an appropriate water-soluble salt, in the presence of cyclodextrin or tromethamine (if needed) in mildly acidified purified water (if stability permits) with or without benzalkonium chloride and polyvinyl alcohol. Amide prodrugs met with mixed success due to incomplete intraocular hydrolysis. Suspension and ocular inserts appear irritating to the inflamed eye. Oil drop may be a suitable option for insoluble drugs and ointment may be used for sustained effect. Recent studies showed that the use of colloidal nanoparticle formulations and the potent COX 2 inhibitor bromfenac may enhance NSAID efficacy in eye preparations.
anti-inflammatory; cyclodextrins; formulation; NSAID; ocular; PGE2
The effect of a homologue series of nonionic surfactants, namely poly(ethylene glycol) (PEG) fatty acid esters, differing in oxyethylene (PEG 8, PEG 12, and PEG 40) and fatty acid (stearate, mono and di-laurate, and mono and di-oleate) chain lengths, on in vitro skin permeability of ketoprofen (KTP) vehicled in plasters was investigated. The drug diffusion through hairless mouse skin as well as the effect of the surfactant type and strength was studied by Franz diffusion cells and ATR-FTIR spectroscopy. The use of PEG stearate series revealed that the surfactant with the largest polar head, namely PEG 40, was ineffective in enhancing the skin permeation of KTP, independently of the plaster concentrations. The effect of the hydrophobic chain was investigated only by using the shortest oxyethylene chains. The experimental results revealed that the oxyethylene chain length of surfactants appeared to be more influent than the alkyl chain. The prediction of the absorption enhancing capability of these PEG derivatives appeared related to the vehicle other than the proper combination of the number of ethylene oxide groups and alkyl groups.
enhancer; hairless mouse skin; ketoprofen; plaster; polyethylene glycol derivatives
The aim of this study was to develop an effective drug delivery system for the simultaneous topical delivery of two anti-inflammatory drugs, spantide II (SP) and ketoprofen (KP). To achieve this primary goal we have developed a skin permeating nanogel system (SPN) containing surface modified polymeric bilayered nanoparticles along with a gelling agent. Poly-(lactide-co-glycolic acid) and chitosan were used to prepare bilayered nanoparticles (NPS) and the surface was modified with oleic acid (NPSO). Hydroxypropyl methyl cellulose (HPMC) and Carbopol with the desired viscosity were utilized to prepare the nanogels. The nanogel system was further investigated for in vitro skin permeation, drug release and stability studies. Allergic contact dermatitis (ACD) and psoriatic plaque like model were used to assess the effectiveness of SPN. Dispersion of NPSO in HPMC (SPN) produced a stable and uniform dispersion. In vitro permeation studies revealed increase in deposition of SP for the SP-SPN or SP+KP-SPN in the epidermis and dermis by 8.5 and 9.5 folds, respectively than SP-gel. Further, the deposition of KP for KP-SPN or SP+KP-SPN in epidermis and dermis was 9.75 and 11.55 folds higher, respectively than KP-gel. Similarly the amount of KP permeated for KP-SPN or SP+KP-SPN was increased by 9.92 folds than KP-gel. The ear thickness in ACD model and the expression of IL-17 and IL-23; PASI score and TEWL values in psoriatic plaque like model were significantly less (p<0.001) for SPN compared to control gel. Our results suggest that SP+KP-SPN have significant potential for the percutaneous delivery of SP and KP to the deeper skin layers for treatment of various skin inflammatory disorders.
Rubbing a topical NSAID (non steroidal anti-inflammatory drug) on the skin may increase local drug permeation, affecting its distribution to the site of pain and inflammation. The present study evaluates this hypothesis, by assessing in vitro the effect on skin permeation of applying diclofenac-dieythylamine 1.16% gel with or without rubbing.
A single dose of 5 mg/cm2 diclofenac-diethylamine 1.16% gel was applied on excised human skin mounted in Franz-type diffusion cells without or with rubbing for 45 s. Drug penetration into the skin layers was determined after 1 h using the tape stripping technique. In vitro cutaneous permeation into the receptor fluid of the diffusion chamber was measured up to 24 h. Skin electrical resistance was also recorded.
Application of diclofenac-diethylamine 1.16% gel with rubbing resulted to a 5-fold higher flux of diclofenac through the skin than when applied without rubbing at 8 h (P = 0.04). Skin rubbing for 45 s decreased by 2-fold skin electrical resistance when compared to the standard application. Application of diclofenac-diethylamine 1.16% gel with rubbing tended to result in higher accumulation in the stripped skin vs. the superficial skin layers when applied without rubbing (P = 0.2).
These results suggest that rubbing may alter the superficial skin layer resulting in a transient faster initial diffusion of topically applied diclofenac through the stratum corneum into the deeper skin layer of the dermis to the tissue target.
Nonsteroidal anti-inflammatory drugs (NSAID) represent a one of the most widely used anti-inflammatory substances. Their anti-inflammatory effects are mainly based on inhibition of cyclooxygenase. The potential direct effect of NSAID on leukocyte migration was poorly investigated. Using time-lapse microscopy and 96-well fluorescence-based assay, we studied the effect of three different NSAID, ketoprofen, diclofenac and SC-560, on leukocyte haptokinesis and haptotaxis in vivo and in vitro.
NSAID induced an immediate inhibiting effect on leukocyte migration both in vitro and in vivo. This effect was dose-dependent and was not restricted to a specific type of leukocytes. The inhibition of leukocyte migration by NSAID was partially re-stored after removal of inhibiting agent. Only complete blockade of leukocyte migration was accompanied by a strong reduction of [Ca2+]i.
NSAID strongly supress leukocyte migration. The results of the present study may have important clinical implications since blockade of leukocyte migration can be achieved after topical application of NSAID.
Acid sphingomyelinase (ASM) is an important early responder in inflammatory cytokine signaling. The role of ASM in retinal vascular inflammation and vessel loss associated with diabetic retinopathy is not known and represents the goal of this study.
RESEARCH DESIGN AND METHODS
Protein and gene expression profiles were determined by quantitative RT-PCR and Western blot. ASM activity was determined using Amplex Red sphingomyelinase assay. Caveolar lipid composition was analyzed by nano-electrospray ionization tandem mass spectrometry. Streptozotocin-induced diabetes and retinal ischemia-reperfusion models were used in in vivo studies.
We identify endothelial caveolae-associated ASM as an essential component in mediating inflammation and vascular pathology in in vivo and in vitro models of diabetic retinopathy. Human retinal endothelial cells (HREC), in contrast with glial and epithelial cells, express the plasma membrane form of ASM that overlaps with caveolin-1. Treatment of HREC with docosahexaenoic acid (DHA) specifically reduces expression of the caveolae-associated ASM, prevents a tumor necrosis factor-α–induced increase in the ceramide-to-sphingomyelin ratio in the caveolae, and inhibits cytokine-induced inflammatory signaling. ASM is expressed in both vascular and neuroretina; however, only vascular ASM is specifically increased in the retinas of animal models at the vasodegenerative phase of diabetic retinopathy. The absence of ASM in ASM−/− mice or inhibition of ASM activity by DHA prevents acellular capillary formation.
This is the first study demonstrating activation of ASM in the retinal vasculature of diabetic retinopathy animal models. Inhibition of ASM could be further explored as a potential therapeutic strategy in treating diabetic retinopathy.
The design and the synthesis of prodrugs for nonsteroidal anti-inflammatory drugs (NSAIDs) have been given much attention by medicinal chemists, especially in the last decade. As a therapeutic group, NSAIDs are among the most widely used prescribed and over the counter (OTC) medications. The rich literature about potential NSAID prodrugs clearly shows a shift from alkyl, aryalkyl or aryl esters with the sole role of masking the carboxylic acid group, to more elaborate conjugates that contain carefully chosen groups to serve specific purposes, such as enhancement of water solubility and dissolution, nitric oxide release, hydrogen sulfide release, antioxidant activity, anticholinergic and acetylcholinesterase inhibitory (AChEI) activity and site-specific targeting and delivery. This review will focus on NSAID prodrugs that have been designed or were, later, found to possess intrinsic pharmacological activity as an intact chemical entity. Such intrinsic activity might augment the anti-inflammatory activity of the NSAID, reduce its side effects or transform the potential therapeutic use from classical anti-inflammatory action to something else. Reports discussed in this review will be those of NO-NSAIDs, anticholinergic and AChEI-NSAIDs, Phospho-NSAIDs and some miscellaneous agents. In most cases, this review will cover literature dealing with these NSAID prodrugs from the year 2006 and later. Older literature will be used when necessary, e.g., to explain the chemical and biological mechanisms of action.
anti-inflammatory; cyclooxygenase; codrug; mutual prodrug; NO-NSAIDs; NSAIDs; Phospho-NSAIDs; prodrug
Airway smooth muscle (ASM) cells have been reported to contribute to the inflammation of asthma. Because the thiazolidinediones (TZDs) exert anti-inflammatory effects, we examined the effects of troglitazone and rosiglitazone on the release of inflammatory moieties from cultured human ASM cells. Troglitazone dose-dependently reduced the IL-1β–induced release of IL-6 and vascular endothelial growth factor, the TNF-α–induced release of eotaxin and regulated on activation, normal T expressed and secreted (RANTES), and the IL-4–induced release of eotaxin. Rosiglitazone also inhibited the TNF-α–stimulated release of RANTES. Although TZDs are known to activate peroxisome proliferator–activated receptor-γ (PPARγ), these anti-inflammatory effects were not affected by a specific PPARγ inhibitor (GW 9662) or by the knockdown of PPARγ using short hairpin RNA. Troglitazone and rosiglitazone each caused the activation of adenosine monophosphate-activated protein kinase (AMPK), as detected by Western blotting using a phospho-AMPK antibody. The anti-inflammatory effects of TZDs were largely mimicked by the AMPK activators, 5-amino-4-imidazolecarboxamide ribose (AICAR) and metformin. However, the AMPK inhibitors, Ara A and Compound C, were not effective in preventing the anti-inflammatory effects of troglitazone or rosiglitzone, suggesting that the effects of these TZDs are likely not mediated through the activation of AMPK. These data indicate that TZDs inhibit the release of a variety of inflammatory mediators from human ASM cells, suggesting that they may be useful in the treatment of asthma, and the data also indicate that the effects of TZDs are not mediated by PPARγ or AMPK.
shRNA; anti-inflammatory; PPARγ; IL-1β; TNF-α
Disruption of the lymphatic vasculature causes edema, inflammation, and end-tissue destruction. To assess the therapeutic efficacy of systemic anti-inflammatory therapy in this disease, we examined the impact of a nonsteroidal anti-inflammatory drug (NSAID), ketoprofen, and of a soluble TNF-α receptor (sTNF-R1) upon tumor necrosis factor (TNF)-α activity in a mouse model of acquired lymphedema.
Methods and Findings
Lymphedema was induced by microsurgical ablation of major lymphatic conduits in the murine tail. Untreated control mice with lymphedema developed significant edema and extensive histopathological inflammation compared to sham surgical controls. Short-term ketoprofen treatment reduced tail edema and normalized the histopathology while paradoxically increasing TNF-α gene expression and cytokine levels. Conversely, sTNF-R1 treatment increased tail volume, exacerbated the histopathology, and decreased TNF-α gene expression. Expression of vascular endothelial growth factor-C (VEGF-C), which stimulates lymphangiogenesis, closely correlated with TNF-α expression.
Ketoprofen therapy reduces experimental post-surgical lymphedema, yet direct TNF-α inhibition does not. Reducing inflammation while preserving TNF-α activity appears to optimize the repair response. It is possible that the observed favorable responses, at least in part, are mediated through enhanced VEGF-C signaling.
The transdermal application of substances represents an elegant approach to overcome side effects related to injections or oral treatment. Due to benefits like a constant plasma level, no pain during application and a simple therapeutic regime, the optimization of formulations for transdermal drug delivery has gained interest in the last decades. Ibuprofen is a non-steroidal anti-inflammatory compound which is nowadays often used transdermally. The objective of this work was to conduct a study on the effect of different 5% ibuprofen containing formulations (Ibutop® cream, Ibutop® gel, and ibuprofen solution in phosphate buffered saline) on the in vitro-percutaneous permeation of ibuprofen through skin to emphasise the importance of the formulation on percutaneous permeation and skin reservoir.
The permeation experiments were conducted in Franz-type diffusion cells according to OECD guideline 428 with 2 mg/cm2 ibuprofen formulation on each skin sample. Ibuprofen was analysed in the receptor fluid and extracted skin samples by UV-VIS high-performance liquid-chromatography at 238 nm. The plot of the cumulative amount of ibuprofen permeated versus time was employed to calculate the apparent permeability coefficient, the maximum flux and the lagtime, all of which were statistically analysed by One-way ANOVA.
Although ibuprofen permeation out of the gel increases rapidly within the first four hours, the cream produced the highest ibuprofen delivery through the skin within 28 hours, followed by the solution and the gel. A significant shorter lagtime was found after gel treatment compared with the cream and the solution. After 28 hours 59% of the applied ibuprofen was found in the receptor fluid of the cream treated samples, 26% in the solution treated samples and 21% in the samples treated with the gel. Fourfold higher ibuprofen reservoirs were found in the solution and gel treated skin samples compared to the cream treated skin samples.
The present study demonstrates the importance of the formulation on transdermal drug delivery of ibuprofen and emphasises the differences of drug storage within the skin due to the formulation. Thus, it is a mistaken assumption that formulations comprising the same drug amount are equivalent regarding skin permeability.
The purpose of this research was the preparation of four formulations containing hydrocortisone acetate (HCA) for topical application, including two aqueous systems (hydrophilic microemulsion and aqueous gel) and two systems with dominant hydrophobicity (hydrophobic microemulsion and ointment). The formulations were tested for the release and permeation of HCA across an animal membrane. The release of HCA was found comparable for the four systems. The two microemulsions promote permeation across an ex-vivo membrane, examined by means of a Franz cell. Hydrophobic microemulsion guarantees the highest solubility (2,370 μg/ml) and flux (133 μg/cm2.h) of the drug, since it contains almost 40% Transcutol, a permeation enhancer. Gel and ointment provide lower solubility and flux, being the values, related to the ointment, the lowest ones (562 μg/ml and 0.4 μg/cm2.h). Experimental results allow the conclusion that gel and ointment can be suitable when it is desirable to minimize absorption of topically applied HCA as to keep the drug restricted to the diseased area and prevent side effects of the systemic presence of HCA.
gel and ointment; hydrocortisone acetate; microemulsions; transdermal permeability
BACKGROUND: Airway smooth muscle (ASM) is considered to be a target for mediators released during airway inflammation. AIMS: To investigate the expression of c-fos, a constituent of the transcription factor activator protein-1, in human ASM cells. In addition, to measure the release of interleukin (IL)-6 into the conditioned medium of stimulated ASM cells, as well as DNA biosynthesis and changes in cell number. METHODS: Serum-deprived human ASM cells in the G0/G1 phase were stimulated with the pro-inflammatory cytokines; tumour necrosis factor-alpha, IL-1beta, IL-5 and IL-6. The expression of mRNA encoding the proto-oncogene c-fos was measured by Northern blot analysis. Cell proliferation was assessed by [3H]-thymidine incorporation assays and cell counting, and IL-6 levels in cell-conditioned medium were measured by enzyme-linked immunosorbent assay. RESULTS: All of the cytokines investigated induced a rapid (within 1 h) and transient increase in the expression of mRNA encoding c-fos, followed by the expression and enhanced release of IL-6. Cell proliferation remained unchanged in cytokine-stimulated cells. CONCLUSIONS: Cytokine-induced c-fos expression in human ASM cells could be described as a marker of cell 'activation'. The possible association of these results with airway inflammation, through secondary intracellular mechanisms such as cytokine production, is discussed.
In asthma, the increase in airway smooth muscle (ASM) can contribute to inflammation, airway wall remodeling and airway hyperresponsiveness (AHR). Targetting peroxisome proliferator-activated receptor γ (PPARγ), a receptor upregulated in ASM in asthmatic airways, may provide a novel approach to regulate these contributions. This review summarises experimental evidence that PPARγ ligands, such as rosiglitazone (RGZ) and pioglitazone (PGZ), inhibit proliferation and inflammatory cytokine production from ASM in vitro. In addition, inhaled administration of these ligands reduces inflammatory cell infiltration and airway remodelling in mouse models of allergen-induced airways disease. PPARγ ligands can also regulate ASM contractility, with acute treatment eliciting relaxation of mouse trachea in vitro through a PPARγ-independent mechanism. Chronic treatment can protect against the loss of bronchodilator sensitivity to β2-adrenoceptor agonists and inhibit the development of AHR associated with exposure to nicotine in utero or following allergen challenge. Of particular interest, a small clinical trial has shown that oral RGZ treatment improves lung function in smokers with asthma, a group that is generally unresponsive to conventional steroid treatment. These combined findings support further investigation of the potential for PPARγ agonists to target the noncontractile and contractile functions of ASM to improve outcomes for patients with poorly controlled asthma.
During recent years, there has been growing interest in use of topical vehicle systems to assist in drug permeation through the skin. Drugs of interest are usually those that are problematic when given orally, such as piroxicam, a highly effective anti-inflammatory, anti-pyretic, and analgesic, but with the adverse effect of causing gastrointestinal ulcers. The present study investigated the in vitro and in vivo pharmacodynamic activity of a newly synthesized palm oil esters (POEs)-based nanocream containing piroxicam for topical delivery.
A ratio of 25:37:38 of POEs: external phase: surfactants (Tween 80:Span 20, in a ratio 80:20), respectively was selected as the basic composition for the production of a nanocream with ideal properties. Various nanocreams were prepared using phosphate-buffered saline as the external phase at three different pH values. The abilities of these formulae to deliver piroxicam were assessed in vitro using a Franz diffusion cell fitted with a cellulose acetate membrane and full thickness rat skin. These formulae were also evaluated in vivo by comparing their anti-inflammatory and analgesic activities with those of the currently marketed gel.
After eight hours, nearly 100% of drug was transferred through the artificial membrane from the prepared formula F3 (phosphate-buffered saline at pH 7.4 as the external phase) and the marketed gel. The steady-state flux through rat skin of all formulae tested was higher than that of the marketed gel. Pharmacodynamically, nanocream formula F3 exhibited the highest anti- inflammatory and analgesic effects as compared with the other formulae.
The nanocream containing the newly synthesized POEs was successful for trans-dermal delivery of piroxicam.
piroxicam; nanocream; analgesic; anti-inflammatory; skin permeation
The initial bronchoconstrictor response of the asthmatic airway depends on airway smooth muscle (ASM) contraction. Intracellular calcium is a key signaling molecule, mediating a number of responses, including proliferation, gene expression, and contraction of ASM. Ca2+ influx through receptor-operated calcium (ROC) or store-operated calcium (SOC) channels is believed to mediate longer term signals. The mechanisms of SOC activation in ASM remain to be elucidated. Recent literature has identified the STIM and ORAI proteins as key signaling players in the activation of the SOC subtype; calcium release–activated channel current (ICRAC) in a number of inflammatory cell types. However, the role for these proteins in activation of SOC in smooth muscle is unclear. We have previously demonstrated a role for STIM1 in SOC channel activation in human ASM. The aim of this study was to investigate the expression and define the potential roles of the ORAI proteins in SOC-associated Ca2+ influx in human ASM cells. Here we show that knockdown of ORAI1 by siRNA resulted in reduced thapsigargin- or cyclopiazonic acid (CPA)–induced Ca2+ influx, without affecting Ca2+ release from stores or basal levels. CPA-induced inward currents were also reduced in the ORAI1 knockdown cells. We propose that ORAI1 together with STIM1 are important contributors to SOC entry in ASM cells. These data extend the major tissue types in which these proteins appear to be major determinants of SOC influx, and suggest that modulation of these pathways may prove useful in the treatment of bronchoconstriction.
airway smooth muscle; ORAI; store-operated calcium entry; ion channels
Ophthalmic bromfenac sodium sesquihydrate is a topically applied selective cyclooxygenase (COX)-2 inhibitor. It is similar to amfenac, except for a bromine atom at the C4 of the benzoyl ring position, which markedly affects its in vitro and in vivo potency, extends the duration of anti-inflammatory activity, and enhances its inhibitory effect on COX-2 absorption across the cornea and penetration into ocular tissues. The United States Food and Drug Administration approved bromfenac in 2005 for the treatment of postoperative inflammation and the reduction of ocular pain in patients who have undergone cataract surgery. Nonsteroidal anti-inflammatory drugs (NSAIDs), and among them bromfenac, could be even more effective than steroids at reestablishing the blood–aqueous barrier, as revealed by flare on slit-lamp examination and as quantitatively measured using ocular fluorophotometry. Similar to other NSAIDs, it has a role in inhibiting intraoperative miosis during cataract surgery. However, bromfenac also seems to be useful in other situations, such as refractive surgery, allergic conjunctivitis (not useful in dry eye), choroidal neovascularization, and even ocular oncology. No reports of systemic toxicity have been published and bromfenac has good topical tolerance with a low incidence of adverse effects.
bromfenac; ophthalmic nonsteroidal anti-inflammatory drugs; inflammation; cataract surgery
Nonsteroidal anti-inflammatory drugs (NSAIDs) induce gastric injury on long-term usage. This study aims at reducing the side effect of NSAIDs by encapsulating in zein, an acid-resistant biopolymer. Aceclofenac-loaded zein microspheres were prepared by emulsification and solvent evaporation method. The stability of zein microspheres at gastric pH retarded the release of the entrapped drug and hence reduces the possibility of gastric injury. However, the in vitro release of aceclofenac was sustained up to 72 h at intestinal pH. Thus, zein microspheres pave the way for the development of safe and sustained delivery system for NSAIDs thereby achieving the desired therapeutic potential with reduced side effects for chronic inflammatory disorders.
biopolymers; drug delivery; gastric injury; NSAIDs; zein microspheres
We investigated the correlation between an in vivo isobologram based on the concentrations of new quinolones (NQs) in brain tissue and the administration of nonsteroidal anti-inflammatory drugs (NSAIDs) for the occurrence of convulsions in mice and an in vitro isobologram based on the concentrations of both drugs for changes in the γ-aminobutyric acid (GABA)-induced current response in Xenopus oocytes injected with mRNA from mouse brains in the presence of NQs and/or NSAIDs. After the administration of enoxacin (ENX) in the presence or absence of felbinac (FLB), ketoprofen (KTP), or flurbiprofen (FRP), a synergistic effect was observed in the isobologram based on the threshold concentration in brain tissue between mice with convulsions and those without convulsions. The three NSAIDs did not affect the pharmacokinetic behavior of ENX in the brain. However, the ENX-induced inhibition of the GABA response in the GABAA receptor expressed in Xenopus oocytes was enhanced in the presence of the three NSAIDs. The inhibition ratio profiles of the GABA responses for both drugs were analyzed with a newly developed toxicodynamic model. The inhibitory profiles for ENX in the presence of NSAIDs followed the order KTP (1.2 μM) > FRP (0.3 μM) > FLB (0.2 μM). These were 50- to 280-fold smaller than those observed in the absence of NSAIDs. The inhibition ratio (0.01 to 0.02) of the GABAA receptor in the presence of both drugs was well-fitted to the isobologram based on threshold concentrations of both drugs in brain tissue between mice with convulsions and those without convulsions, despite the presence of NSAIDs. In mice with convulsions, the inhibitory profiles of the threshold concentrations of both drugs in brain tissue of mice with convulsions and those without convulsions can be predicted quantitatively by using in vitro GABA response data and toxicodynamic model.
Excessive airway obstruction is the cause of symptoms and abnormal lung function in asthma.
As airway smooth muscle (ASM) is the effecter controlling airway calibre, it is suspected that dysfunction of ASM contributes to the pathophysiology of asthma. However, the precise role of ASM in the series of events leading to asthmatic symptoms is not clear. It is not certain whether, in asthma, there is a change in the intrinsic properties of ASM, a change in the structure and mechanical properties of the noncontractile components of the airway wall, or a change in the interdependence of the airway wall with the surrounding lung parenchyma. All these potential changes could result from acute or chronic airway inflammation and associated tissue repair and remodelling.
Anti-inflammatory therapy, however, does not “cure” asthma, and airway hyperresponsiveness can persist in asthmatics, even in the absence of airway inflammation. This is perhaps because the therapy does not directly address a fundamental abnormality of asthma, that of exaggerated airway narrowing due to excessive shortening of ASM.
In the present study, a central role for airway smooth muscle in the pathogenesis of airway hyperresponsiveness in asthma is explored.
Airway mechanics; interdependence; lung function; muscle adaptation; muscle contraction; parenchyma
The nonsteroidal anti-inflammatory drug (NSAID), indomethacin (Indo), has a large number of divergent biological effects, the molecular mechanism(s) for which have yet to be fully elucidated. Interestingly, Indo is highly amphiphilic and associates strongly with lipid membranes, which influence localization, structure and function of membrane-associating proteins and actively regulate cell signaling events. Thus, it is possible that Indo regulates diverse cell functions by altering micro-environments within the membrane. Here we explored the effect of Indo on the nature of the segregated domains in a mixed model membrane composed of dipalmitoyl phosphatidyl-choline (di16∶0 PC, or DPPC) and dioleoyl phosphatidyl-choline (di18∶1 PC or DOPC) and cholesterol that mimics biomembranes.
Using a series of fluorescent probes in a fluorescence resonance energy transfer (FRET) study, we found that Indo induced separation between gel domains and fluid domains in the mixed model membrane, possibly by enhancing the formation of gel-phase domains. This effect originated from the ability of Indo to specifically target the ordered domains in the mixed membrane. These findings were further confirmed by measuring the ability of Indo to affect the fluidity-dependent fluorescence quenching and the level of detergent resistance of membranes.
Because the tested lipids are the main lipid constituents in cell membranes, the observed formation of gel phase domains induced by Indo potentially occurs in biomembranes. This marked Indo-induced change in phase behavior potentially alters membrane protein functions, which contribute to the wide variety of biological activities of Indo and other NSAIDs.
Acid sphingomyelinase (ASM) hydrolyses sphingomyelin and generates the lipid messenger ceramide, which mediates a variety of stress-related cellular processes. The pathological effects of dysregulated ASM activity are evident in several human diseases and indicate an important functional role for ASM regulation. We investigated alternative splicing as a possible mechanism for regulating cellular ASM activity.
We identified three novel ASM splice variants in human cells, termed ASM-5, -6 and -7, which lack portions of the catalytic- and/or carboxy-terminal domains in comparison to full-length ASM-1. Differential expression patterns in primary blood cells indicated that ASM splicing might be subject to regulatory processes. The newly identified ASM splice variants were catalytically inactive in biochemical in vitro assays, but they decreased the relative cellular ceramide content in overexpression studies and exerted a dominant-negative effect on ASM activity in physiological cell models.
These findings indicate that alternative splicing of ASM is of functional significance for the cellular stress response, possibly representing a mechanism for maintaining constant levels of cellular ASM enzyme activity.