Local application of neurotensin to abdominal organs triggers cardiovascular reflexes in guinea pigs: possible mechanisms

Intraabdominal (IAB) injections or topical application of neurotensin (NT) to the serosal surface of the ileum or stomach evoked dose-dependent increases of blood pressure and of heart rate in anesthetized guinea pigs. These effects were markedly reduced by prior animal treatment with a ganglion blocker, alpha and beta adrenoceptor blockers, as well as by exposure of the abdominal organs to lidocaine, a local anesthetic. The blood pressure and heart rate responses to IAB injections or topical application of NT to the ileum or stomach were both inhibited by animal pretreatment with capsaicin. Cervical vagotomy or atropine pretreatment did not prevent or alter the cardiovascular responses to IAB injections of NT. These results suggest the presence in some organs and/or tissues of the abdominal cavity of sympathetic, capsaicin-sensitive sensory nerve fibers which, upon stimulation by NT, produce reflex increases of blood pressure and of heart rate.     

SYSTEMIC TOXIC REACTIONS TO LOCAL ANESTHETICS

The topical use of anesthetic agents involves an element of risk. Systemic toxic reactions are rare, but they do occur and may result in death. When a reaction occurs from a topical application, it usually progresses rapidly to respiratory and cardiovascular collapse, and thus therapy must be instituted with more haste to avoid deaths. Fatal systemic toxic reactions from topically administered anesthetic drugs are, in effect, usually not due to well informed use of the drug but to misuse owing to less than complete understanding of absorption.Emphasis is placed on the causes, prophylaxis and treatment of severe systemic toxic reactions which follow the topical application of local anesthetic drugs. If systemic toxic reactions resulting from a safe dose of a local anesthetic agent are correctly treated, there will usually follow an uneventful recovery rather than a catastrophe.     

Analysis of Chinese herbal creams prescribed for dermatological conditions

ObjectiveTo determine whether Chinese herbal creams used for the treatment of dermatological conditions contain steroids.Design11 herbal creams obtained from patients attending general and paediatric dermatology outpatient clinics were analysed with high resolution gas chromatography and mass spectrometry. SettingDepartments of dermatology and clinical biochemistry.ResultsEight creams contained dexamethasone at a mean concentration of 456 μg/g (range 64 to 1500 μg/g). All were applied to areas of sensitive skin such as face and flexures.ConclusionGreater regulation needs to be imposed on Chinese herbalists to prevent illegal and inappropriate prescribing of potent steroids.

Key messages

• Patients with eczema often report improvement with Chinese herbal creams
• There may be no indication on the label about the contents of the cream
• Eight of the 11 creams analysed contained dexamethasone at concentrations inappropriate for use on the face or in children
• Inadvertent use of topical steroids can cause severe exacerbation of eczema herpeticum
• Closer regulation of herbal medicines is required

     

Patients’ view of the differences in topical creams for radiation dermatitis prevention. A pilot study of cosmetic properties

AimTo investigate the feasibility of including patients’ reports on the cosmetic properties of topical formulations for acute radiation dermatitis (ARD).BackgroundNo topical agent tested for acute radiation dermatitis (ARD) has proven to be better than any other, all achieving similar objective outcomes. No clear guidelines have therefore been established in clinics. Because the vehicle for such creams has shown to be an important factor in patient adherence to treatments in other dermatological diseases, patients’ opinions are evaluated.Material and methodsSeventy breast cancer patients referred for postoperative radiotherapy after conservative surgery were enrolled. Patients were assigned to use one of the 7 topical agents that are most-commonly used in the prevention of ARD. Patients’ reports were assessed using continuous visual analogue scales (VAS), objective signs and symptoms produced by ARD, and were rated using the RTOG and RISRAS scales.ResultsThe creams tested differed in their cosmetic properties significantly (p = 0.044). The performance of the agent, their absorption and any residue left over were also significantly different (p = 0.022, 0.014 and 0.02, respectively).ConclusionsTopical agents for preventive ARD are reported by patients to show different cosmetic properties. Cosmetic properties are important when choosing topical agents for ARD prevention. Recommending those with better cosmetic profiles would improve patient adherence to treatments.     

Viscoelastic evaluation of topical creams containing microcrystalline cellulose/sodium carboxymethyl cellulose as stabilizer

The purpose of this study was to examine the viscoelastic properties of topical creams containing various concentrations of microcrystalline cellulose and sodium carboxymethyl cellulose (Avicel(R) CL-611) as a stabilizer. Avicel CL-611 was used at 4 different levels (1%, 2%, 4%, and 6% dispersion) to prepare topical creams, and hydrocortisone acetate was used as a model drug. The viscoelastic properties such as loss modulus (G"), storage modulus (G'), and loss tangent (tan delta) of these creams were measured using a TA Instruments AR 1000 Rheometer and compared to a commercially available formulation. Continuous flow test to determine the yield stress and thixotropic behavior, and dynamic mechanical tests for determining the linear viscosity time sweep data, were performed. Drug release from the various formulations was studied using an Enhancer TM Cell assembly. Formulations containing 1% and 2% Avicel CL-611 had relative viscosity, yield stress, and thixotropic values that were similar to those of the commercial formulation. The elastic modulus (G') of the 1% and 2% formulation was relatively high and did not cross the loss modulus (G"), indicating that the gels were strong. In the commercial formulation, G' increased after preshearing and broke down after 600 seconds. The strain sweep tests showed that for all formulations containing Avicel CL-611, the G' was above G" with a good distance between them. The gel strength and the predominance of G' can be ranked 6% > 4% > 2%. The strain profiles for the 1% and 2% formulations were similar to those of the commercial formulation. The delta values for the 1% and 2% formulations were similar, and the formulations containing 4% Avicel CL-611 had lower delta values, indicating greater elasticity. Drug release from the commercial preparation was fastest compared to the formulations prepared using Avicel CL-611, a correlation with the viscoelastic properties. It was found that viscoelastic data, especially the strain sweep profiles of products containing Avicel CL-611 1% and 2%, correlated with the commercial formulation. Rheological tests that measure the viscosity, yield stress, thixotropic behavior, other oscillatory parameters such as G' and G" are necessary tools in predicting performance of semisolids.     

Physiology and behavior of pigs before and after castration: effects of two topical anesthetics

Surgical castration of male piglets is a common management practice conducted on commercial swine farms to prevent the occurrence of boar taint and aggressive behavior. However, the procedure of surgical castration causes acute pain-induced distress, which is an animal welfare concern. The objective of this study was to evaluate the use of two topical anesthetics to alleviate the pain caused by castration in piglets as measured by physiological and behavior indices of stress. At 3 days of age, 40 weight-matched piglets were allocated to one of four treatment groups. Treatments included: (i) sham castration (CON), (ii) surgical castration (CAS), (iii) castration and short-acting local anesthetic applied topically to the castration wound (SHORT) and (iv) castration and long-acting local anesthetic applied topically to the castration wound (LONG). Blood samples were collected from piglets before and 30, 60, 120 and 180 min after castration to measure leukocyte and differential counts and cortisol concentrations. The above experiment was repeated without blood collection and behavior was recorded for 30 min before and 180 min after castration or handling. Stress vocalizations were recorded during castration and handling. All piglets were weighed before and 24 h after castration and wound healing was recorded daily for the first 14 days after castration. Leukocyte counts and differentials did not differ (P > 0.05) among any of the treatments. Cortisol concentrations were elevated (P < 0.06) in CAS, SHORT and LONG piglets compared with controls 30 and 60 min after castration. The percentage of stress vocalizations was greater (P < 0.05) among castrated piglets compared with CON piglets, regardless of anesthetic treatment. Piglets that were castrated and not given a topical anesthetic spent more time (P < 0.05) lying without contact compared with piglets castrated and given a topical anesthetic, regardless of the topical anesthetic administered. Body weight change did not differ (P > 0.05) among treatments 24 h after castration or control handling and wound healing scores were greater (P < 0.05) in SHORT compared with CAS and LONG piglets 9 to 14 days after castration. In this study, the use of a short- or long-acting topical anesthetic was not effective in reducing the pain-induced distress caused by castration in piglets. Further research is needed to evaluate alternative practical methods to reduce the pain caused by the on-farm castration of piglets.     

Antihirsutism activity of Fennel (fruits of Foeniculum vulgare) extract. A double-blind placebo controlled study.

Idiopathic hirsutism is defined as the occurrence of excessive male pattern hair growth in women who have a normal ovulatory menstrual cycle and normal levels of serum androgens. It may be a disorder of peripheral androgen metabolism. In this study we evaluated the clinical response of idiopathic hirsutism to topical Fennel extract. Fennel, Foeniculum vulgare, is a plant, which has been used as an estrogenic agent. The ethanolic extract of Fennel was obtained by using a soxhlete apparatus. In a double blind study, 38 patients were treated with creams containing 1%, 2% of Fennel extract and placebo. Hair diameter was measured and rate of growth was considered. The efficacy of treatment with the cream containing 2% Fennel is better than the cream containing 1% Fennel and these two were more potent than placebo. The mean values of hair diameter reduction was 7.8%, 18.3% and -0.5% for patients receiving the creams containing 1%, 2% and 0% (placebo) respectively.     

Topical ocular anesthetics in ocular irritancy testing: a review.

The routine use of topical anesthetics to alleviate discomfort associated with in vivo ocular irritancy testing has been advocated. This review provides information about the adverse effects of topical ocular anesthetics and answers the questions: are topical anesthetics practical and effective in ocular irritancy protocols, is long-term use contraindicated, will topical anesthetics alter the response of a test substance, and are there significant side-effects which might cause pain and suffering in test animals? There was no evidence to support the use of a specific topical anesthetic. Further, information about using systemic analgesics or combinations with local anesthetics that would effectively alleviate discomfort associated with ocular irritancy testing without affecting test results was not found. Comprehensive studies are needed to identify the most effective combination of drugs that would ameliorate discomfort associated with ocular irritation testing.     

Prevention of ocular scarring post glaucoma filtration surgery using the inflammatory cell and platelet binding modulator saratin in a rabbit model

Clinical Relevance

Late complications can occur with use of current antimetabolites to prevent scarring following glaucoma filtration surgery (GFS). Safer, more targeted, anti-fibrosis agents are sought.

Objectives

The protein saratin has been shown to exhibit anti-fibrotic and anti-thrombotic properties in response to injury, but had not been used for glaucoma surgery. The goal of this study was to compare the efficacy of saratin with that of the widely accepted mitomycin-C (MMC) in prolonging bleb survival following GFS in the rabbit model. Two saratin delivery routes were compared; a single intraoperative topical application versus a combination of intraoperative topical application with two additional postoperative injections.

Methods

Twenty-four New Zealand White rabbits underwent GFS and received either intraoperative topical saratin, intraoperative topical saratin plus two injections on post-operative days 4 and 8, balanced saline solution (BSS), or MMC. The bleb tissues and their elevation durations were compared based on clinical and histological findings.

Results

Rabbits receiving topical+injections of saratin had a mean bleb survival of 33.6±8.5 days, significantly higher than the negative BSS controls, which averaged 17.4±6.0 days (p = 0.018). No improvement over BSS was seen for rabbits receiving topical saratin only (15.5±4.8 days, p = 0.749). Rabbits receiving saratin did not develop bleb avascularity and thinning associated with MMC treatment and there were no apparent clinical signs of toxicity.

Conclusions

Treatment with a single intraoperative topical application plus two additional postoperative injections significantly prolonged bleb elevation comparable to MMC, but without toxicity; however, topical application alone was ineffective.     

Liposomal-benzocaine gel formulation: correlation between in vitro assays and in vivo topical anesthesia in volunteers

The aim of the present study was to characterize a liposome-based benzocaine (BZC) formulation designed for topical use on the oral mucosa and to evaluate its in vitro retention and permeation using the Franz-type diffusion cells through pig esophagus mucosa. To predict the effectiveness of new designed formulations during preclinical studies, a correlation between in vitro assays and in vivo efficacy was performed. Liposomal BZC was characterized in terms of membrane/water partition coefficient, encapsulation efficiency, size, polydispersity, zeta potential, and morphology. Liposomal BZC (BL10) was incorporated into gel formulation and its performances were compared to plain BZC gel (B10) and the commercially available BZC gel (B20). BL10 and B10 presented higher flux and retention on pig esophagus mucosa with a shorter lag time, when compared to B20. BZC flux was strongly correlated with in vivo anesthetic efficacy, but not with topical anesthesia duration. The retention studies did not correlate with any of the in vivo efficacy parameters. Thus, in vitro permeation study can be useful to predict anesthetic efficacy during preclinical tests, because a correlation between flux and anesthetic efficacy was observed. Therefore, in vitro assays, followed by in vivo efficacy, are necessary to confirm anesthetic performance.     

Acaricidal Activity of Eugenol Based Compounds against Scabies Mites

Backgound

Human scabies is a debilitating skin disease caused by the “itch mite” Sarcoptes scabiei. Ordinary scabies is commonly treated with topical creams such as permethrin, while crusted scabies is treated with topical creams in combination with oral ivermectin. Recent reports of acaricide tolerance in scabies endemic communities in Northern Australia have prompted efforts to better understand resistance mechanisms and to identify potential new acaricides. In this study, we screened three essential oils and four pure compounds based on eugenol for acaricidal properties.

Methodology/Principal Findings

Contact bioassays were performed using live permethrin-sensitive S. scabiei var suis mites harvested from pigs and permethrin-resistant S. scabiei var canis mites harvested from rabbits. Results of bioassays showed that clove oil was highly toxic against scabies mites. Nutmeg oil had moderate toxicity and ylang ylang oil was the least toxic. Eugenol, a major component of clove oil and its analogues –acetyleugenol and isoeugenol, demonstrated levels of toxicity comparable to benzyl benzoate, the positive control acaricide, killing mites within an hour of contact.

Conclusions

The acaricidal properties demonstrated by eugenol and its analogues show promise as leads for future development of alternative topical acaricides to treat scabies.     

Topical Cream-Based Dosage Forms of the Macrocyclic Drug Delivery Vehicle Cucurbit[6]uril

The macrocycle family of molecules called cucurbit[n]urils are potential drug delivery vehicles as they are able to form host-guest complexes with many different classes of drugs. This study aimed to examine the utility of Cucurbit[6]uril (CB[6]) in topical cream-based formulations for either localised treatment or for transdermal delivery. Cucurbit[6]uril was formulated into both buffered cream aqueous- and oily cream-based dosage forms. The solid state interaction of CB[6] with other excipients was studied by differential scanning calorimetry and the macrocycle''s transdermal permeability was determined using rat skin. Significant solid state interactions were observed between CB[6] and the other dosage form excipients. At concentrations up to 32% w/w the buffered aqueous cream maintained its normal consistency and could be effectively applied to skin, but the oily cream was too stiff and is not suitable as a dosage form. Cucurbit[6]uril does not permeate through skin; as such, the results imply that cucurbituril-based topical creams may potentially only have applications for localised skin treatment and not for transdermal drug delivery.     

Targeted localized use of therapeutic antibodies: a review of non-systemic,topical and oral applications

Therapeutic antibodies provide important tools in the “medicine chest” of today’s clinician for the treatment of a range of disorders. Typically monoclonal or polyclonal antibodies are administered in large doses, either directly or indirectly into the circulation, via a systemic route which is well suited for disseminated ailments. Diseases confined within a specific localized tissue, however, may be treated more effectively and at reduced cost by a delivery system which targets directly the affected area. To explore the advantages of the local administration of antibodies, we reviewed current alternative, non-systemic delivery approaches which are in clinical use, being trialed or developed. These less conventional approaches comprise: (a) local injections, (b) topical and (c) peroral administration routes. Local delivery includes intra-ocular injections into the vitreal humor (i.e. Ranibizumab for age-related macular degeneration), subconjunctival injections (e.g. Bevacizumab for corneal neovascularization), intra-articular joint injections (i.e. anti-TNF alpha antibody for persistent inflammatory monoarthritis) and intratumoral or peritumoral injections (e.g. Ipilimumab for cancer). A range of other strategies, such as the local use of antibacterial antibodies, are also presented. Local injections of antibodies utilize doses which range from 1/10th to 1/100th of the required systemic dose therefore reducing both side-effects and treatment costs. In addition, any therapeutic antibody escaping from the local site of disease into the systemic circulation is immediately diluted within the large blood volume, further lowering the potential for unwanted effects. Needle-free topical application routes become an option when the condition is restricted locally to an external surface. The topical route may potentially be utilized in the form of eye drops for infections or corneal neovascularization or be applied to diseased skin for psoriasis, dermatitis, pyoderma gangrenosum, antibiotic resistant bacterial infections or ulcerated wounds. Diseases confined to the gastrointestinal tract can be targeted directly by applying antibody via the injection-free peroral route. The gastrointestinal tract is unusual in that its natural immuno-tolerant nature ensures the long-term safety of repeatedly ingesting heterologous antiserum or antibody materials. Without the stringent regulatory, purity and clean room requirements of manufacturing parenteral (injectable) antibodies, production costs are minimal, with the potential for more direct low-cost targeting of gastrointestinal diseases, especially with those caused by problematic antibiotic resistant or toxigenic bacteria (e.g. Clostridium difficile, Helicobacter pylori), viruses (e.g. rotavirus, norovirus) or inflammatory bowel disease (e.g. ulcerative colitis, Crohn’s disease). Use of the oral route has previously been hindered by excessive antibody digestion within the gastrointestinal tract; however, this limitation may be overcome by intelligently applying one or more strategies (i.e. decoy proteins, masking therapeutic antibody cleavage sites, pH modulation, enzyme inhibition or encapsulation). These aspects are additionally discussed in this review and novel insights also provided. With the development of new applications via local injections, topical and peroral routes, it is envisaged that an extended range of ailments will increasingly fall within the clinical scope of therapeutic antibodies further expanding this market.     

Chemotherapy of Aspergillus fumigatus keratitis: An experimental study

An experimental Keratitis study of Aspergillus fumigatus was performed in 130 rabbits divided into 12 groups of ten animals each. Three antifungal drugs (myconazole, amphotericin B and pimaricin) were tested using two procedures (topical drops and subconjunctival injections) and two different concentrations (500 and 10 000 times the MIC). In each case, the drugs were applied every 3 h starting 14 h after inoculation. Miconazole was useful at 10 mg/ ml concentration by topical drops and subconjunctival injections, but was less useful at 5 mg/ ml. Amphotericin B was useful at 5 mg/ ml concentration by topical drops and less useful at 2 mg/ ml.No differences were found between the two concentrations by subconjunctival administration. Pimaricin was useful by topical drops at 50 mg/ ml concentration and less useful at 10 mg/ ml as well as by subconjunctival injections.     

Microparticulate Based Topical Delivery System of Clobetasol Propionate

Psoriasis is a chronic, autoimmune skin disease affecting approximately 2% of the world's population. Clobetasol propionate which is a superpotent topical corticosteroid is widely used for topical treatment of psoriasis. Conventional dosage forms like creams and ointments are commonly prefered for the therapy. The purpose of this study was to develop a new topical delivery system in order to provide the prolonged release of clobetasol propionate and to reduce systemic absorption and side effects of the drug. Clobetasol propionate loaded-poly(D,L-lactic-co-glycolic acid) (PLGA) microspheres were prepared by oil-in-water emulsion–solvent evaporation technique. Particle size analysis, morphological characterization, DSC and XRD analyses and in vitro drug release studies were performed on the microparticle formulations. Emulgel formulations were prepared as an alternative for topical delivery of clobetasol propionate. In vitro drug release studies were carried out from the emulgel formulations containing pure drug and drug-loaded microspheres. In addition, the same studies were performed to determine the drug release from the commercial cream product of clobetasol propionate. The release of clobetasol propionate from the emulgel formulations was significantly higher than the commercial product. In addition, the encapsulation of clobetasol propionate in the PLGA microspheres significantly delayed the drug release from the emulgel formulation. As a result, the decrease in the side effects of clobetasol propionate by the formulation containing PLGA microspheres is expected.     

Biobrane versus 1% silver sulfadiazine in second-degree pediatric burns

Partial-thickness burns in children have been treated for many years by daily, painful tubbing, washing, and cleansing of the burn wound, followed by topical application of antimicrobial creams. Pain and impaired wound healing are the main problems. We hypothesized that the treatment of second-degree burns with Biobrane is superior to topical treatment. Twenty pediatric patients were prospectively randomized in two groups to compare the efficacy of Biobrane versus 1% silver sulfadiazine. The rest of the routine clinical protocols were followed in both groups. Demographic data, wound healing time, length of hospital stay, pain assessments and pain medication requirements, and infection were analyzed and compared. Main outcome measures included pain, pain medication requirements, wound healing time, length of hospital stay, and infection. The application of Biobrane to partial-thickness burns proved to be superior to the topical treatment. Patients included in the biosynthetic temporary cover group presented with less pain and required less pain medication. Length of hospital stay and wound healing time were also significantly shorter in the Biobrane group. None of the patients in either group presented with wound infection or needed skin autografting. In conclusion, the treatment of partial-thickness burns with Biobrane is superior to topical therapy with 1% silver sulfadiazine. Pain, pain medication requirements, wound healing time, and length of hospital stay are significantly reduced.     

Silver as biocides in burn and wound dressings and bacterial resistance to silver compounds

Silver products have been used for thousands of years for their beneficial effects, often for hygiene and in more recent years as antimicrobials on wounds from burns, trauma, and diabetic ulcers. Silver sulfadiazine creams (Silvazine and Flamazine) are topical ointments that are marketed globally. In recent years, a range of wound dressings with slow-release Ag compounds have been introduced, including Acticoat, Actisorb Silver, Silverlon, and others. While these are generally accepted as useful for control of bacterial infections (and also against fungi and viruses), key issues remain, including importantly the relative efficacy of different silver products for wound and burn uses and the existence of microbes that are resistant to Ag⁺. These are beneficial products needing further study, although each has drawbacks. The genes (and proteins) involved in bacterial resistance to Ag have been defined and studied in recent years.     

Pharmacokinetic nature of tachyphylaxis to lidocaine: Peripheral nerve blocks and infiltration anesthesia in rats

Tachyphylaxis to peripheral neural blockade was determined with repeated injections of a constant dose of lidocaine in three experimental models: sciatic nerve block, produced by intraneural or extraneural injections, and infiltration anesthesia. A decrease in the duration of the subsequent blocks was used as the index of tachyphylaxis development The anesthetic content in the nerve or skin was determined using radiolabeled lidocaine. Repeated injections of a constant dose of lidocaine resulted in a marked decrease in the duration of the blocks. Accelerated decline in lidocaine content of nerve or skin was observed with repeated blocks. Our data show that tachyphylaxis rapidly develops with both sciatic nerve blocks and infiltration anesthesia. The data also suggest that the mechanism is largely pharmacokinetic in nature.     

The effect of topical instillation of 2.5% phenylephrine and 1.0% tropicamide on the blood pressure of hypertensive patients

Background: Pupillary dilation is necessary to complete a thorough examination of the internal ocular structures and perform threshold visual fields on the automated perimeter. In our clinic, the topical instillation of 2.5% phenylephrine and 1.0% tropicamide following one drop of topical anesthetic is used routinely for pupil dilation. The vasoconstrictive effects of phenylephrine can cause an increase in peripheral resistance resulting in elevation of systolic and diastolic blood pressures. A rise in systemic blood pressure has been shown to occur following topical instillation of phenylephrine (Heath, Arch Ophthalmol, 1936;16:839–846). This study investigates the effect of topical instillation of 2.5% phenylephrine and 1.0% tropicamide on the blood pressure of known hypertensive patients 30 and 70 min after instillation. Methods: 118 hypertensive patients, all of whom were being treated with anti-hypertensive medications, were involved in the study. Fifty-six patients were dilated with two drops 2.5% phenylephrine and two drops 1.0% tropicamide instilled 5 min apart after one drop of local anesthetic (proparacaine 0.5%). The remaining 62 patients were examined but not dilated. Blood pressure was measured using a sphygmomanometer and stethoscope (right arm sitting) prior to dilation and 30 and 70 min following drop instillation. Results: No clinically significant increase in blood pressure at 30 and 70 min after instillation was observed in the hypertensive group that was dilated. In addition, the change in blood pressure of the dilated group and undilated group was not statistically significant. Conclusion: This study shows that pupillary dilation with 2.5% phenylephrine and 1.0% tropicamide did not significantly increase systemic blood pressure in this population of hypertensive patients.     

Cyclooxygenase-2 and adenylate cyclase/protein kinase A signaling pathway enhances angiogenesis through induction of vascular endothelial growth factor in rat sponge implants

Angiogenesis is reportedly enhanced by prostaglandins (PGs). In the present experiment, we tested whether or not COX-2 and adenylate cyclase/protein kinase A (AC/PKA)-dependent VEGF induction enhanced angiogenesis in this model. Angiogenesis was enhanced by topical injection of human recombinant basic fibroblast growth factor (bFGF). The enhanced angiogenesis by bFGF was inhibited by indomethacin or selective COX-2 inhibitors, NS398, nimesulide, and JTE-522. Topical daily injections of 8-bromo-cAMP enhanced angiogenesis in a dose-dependent manner. Forskolin, an activator of AC, also facilitated angiogenesis in a dose-dependent manner, as did amrinone, an inhibitor of phosphodiesterase. VEGF induction was confirmed by the increased levels in the fluids in the sponge matrix after topical injection of 8-bromo-cAMP. Immunohistochemical investigation further revealed the VEGF-expressed cells in the sponge granulation tissues to be fibroblasts, and the intensity of positive reactions was enhanced by bFGF, 8-bromo-cAMP, forskolin, and amrinone. Angiogenesis was inhibited by indometacin or selective COX-2 inhibitors, NS-398, nimesulide, and JTE-522. In addition, angiogenesis without topical injections of the above compounds was also suppressed by SQ22,536, an inhibitor for AC. or H-89, an inhibitor for PKA, with concomitant reductions in VEGF levels. Daily topical injections of neutralizing antibody or anti-sense oligonucleotide against VEGF significantly suppressed angiogenesis. These results suggested that COX-2 and AC/PKA-dependent induction of VEGF certainly enhanced angiogenesis, and that pharmacological tools for controlling this signaling pathway may be able to facilitate the management of conditions involving angiogenesis.