Anti-allergic activities of Umbelliferone against histamine- and Picryl chloride-induced ear edema by targeting Nrf2/iNOS signaling in mice.

BACKGROUND: The current study was aimed to investigate the anti-allergic activities of the Umbelliferone (UMB) against the acute Histamine and chronic Picryl chloride (PiCl)-induced allergy in mice. UMB is a coumarin derivative (isolated from Angelica decursiva) found in various parts of the plants such as flowers, roots and, stems isolated from the plants of Umbelliferae family. METHODS: The UMB (1, 10, 50 mg/kg) was administered intraperitoneally (i.p) half an h before or 2 h after the induction of allergic ear edema. The acute ear edema was induced by histamine (intradermally, i.d), while the chronic ear edema was induced by painting the PiCl (sensitized with the toluene) on the ear. The antioxidants and oxidative stress markers were assessed. The histological changes were assessed using Hematoxylin and eosin (H and E) and giemsa staining. The immunohistochemistry studies were performed to assess the expression of the nuclear factor erythroid 2-related factor 2 (Nrf2) and inducible nitric oxide synthase (iNOS). The data was analyzed using one-way ANOVA tests followed by Tukey's test with p < 0.05 was chosen as criteria for statistical significance. RESULTS: UMB treatment markedly reduced the allergic ear edema and ear weight compared to the negative control. Furthermore, the UMB attenuated the oxidative stress markers, while induced the antioxidants enzymes. Similarly, the UMB treatment significantly attenuated the serum immunoglobulin E (IgE) level. The UMB treatment markedly improved the histological parameters using H and E staining and Giemsa staining. The UMB administration induced the Nrf2 expression, while attenuated the iNOS expression. Furthermore, the computational analysis was performed to assess the interaction of the UMB with the various protein targets and to determine the mechanism of interaction with the target proteins. CONCLUSION: In conclusion, the UMB treatment significantly alleviated the allergic symptoms, attenuating the oxidative stress, improved the histological features using in vivo and computational approaches.

Ethanol treatment of nanoPGA/PCL composite scaffolds enhances human chondrocyte development in the cellular microenvironment of tissue-engineered auricle constructs.

A major obstacle for tissue engineering ear-shaped cartilage is poorly developed tissue comprising cell-scaffold constructs. To address this issue, bioresorbable scaffolds of poly-ε-caprolactone (PCL) and polyglycolic acid nanofibers (nanoPGA) were evaluated using an ethanol treatment step before auricular chondrocyte scaffold seeding, an approach considered to enhance scaffold hydrophilicity and cartilage regeneration. Auricular chondrocytes were isolated from canine ears and human surgical samples discarded during otoplasty, including microtia reconstruction. Canine chondrocytes were seeded onto PCL and nanoPGA sheets either with or without ethanol treatment to examine cellular adhesion in vitro. Human chondrocytes were seeded onto three-dimensional bioresorbable composite scaffolds (PCL with surface coverage of nanoPGA) either with or without ethanol treatment and then implanted into athymic mice for 10 and 20 weeks. On construct retrieval, scanning electron microscopy showed canine auricular chondrocytes seeded onto ethanol-treated scaffolds in vitro developed extended cell processes contacting scaffold surfaces, a result suggesting cell-scaffold adhesion and a favorable microenvironment compared to the same cells with limited processes over untreated scaffolds. Adhesion of canine chondrocytes was statistically significantly greater (p ≤ 0.05) for ethanol-treated compared to untreated scaffold sheets. After implantation for 10 weeks, constructs of human auricular chondrocytes seeded onto ethanol-treated scaffolds were covered with glossy cartilage while constructs consisting of the same cells seeded onto untreated scaffolds revealed sparse connective tissue and cartilage regeneration. Following 10 weeks of implantation, RT-qPCR analyses of chondrocytes grown on ethanol-treated scaffolds showed greater expression levels for several cartilage-related genes compared to cells developed on untreated scaffolds with statistically significantly increased SRY-box transcription factor 5 (SOX5) and decreased interleukin-1α (inflammation-related) expression levels (p ≤ 0.05). Ethanol treatment of scaffolds led to increased cartilage production for 20- compared to 10-week constructs. While hydrophilicity of scaffolds was not assessed directly in the present findings, a possible factor supporting the summary data is that hydrophilicity may be enhanced for ethanol-treated nanoPGA/PCL scaffolds, an effect leading to improvement of chondrocyte adhesion, the cellular microenvironment and cartilage regeneration in tissue-engineered auricle constructs.

A guide to managing disorders of the ear pinna and canal.

This review will help you troubleshoot everything from infections and foreign bodies to trauma and neoplasm.

Epigenetic inhibitor zebularine activates ear pinna wound closure in the mouse.

BACKGROUND: Most studies on regenerative medicine focus on cell-based therapies and transplantations. Small-molecule therapeutics, though proved effective in different medical conditions, have not been extensively investigated in regenerative research. It is known that healing potential decreases with development and developmental changes are driven by epigenetic mechanisms, which suggests epigenetic repression of regenerative capacity. METHODS: We applied zebularine, a nucleoside inhibitor of DNA methyltransferases, to stimulate the regenerative response in a model of ear pinna injury in mice. FINDINGS: We observed the regeneration of complex tissue that was manifested as improved ear hole repair in mice that received intraperitoneal injections of zebularine. Six weeks after injury, the mean hole area decreased by 83.2 ±â€¯9.4% in zebularine-treated and by 43.6 ±â€¯15.4% in control mice (p < 10-30). Combined delivery of zebularine and retinoic acid potentiated and accelerated this effect, resulting in complete ear hole closure within three weeks after injury. We found a decrease in DNA methylation and transcriptional activation of neurodevelopmental and pluripotency genes in the regenerating tissues. INTERPRETATION: This study is the first to demonstrate an effective induction of complex tissue regeneration in adult mammals using zebularine. We showed that the synergistic action of an epigenetic drug (zebularine) and a transcriptional activator (retinoic acid) could be effectively utilized to induce the regenerative response, thus delineating a novel pharmacological strategy for regeneration. The strategy was effective in the model of ear pinna regeneration in mice, but zebularine acts on different cell types, therefore, a similar approach can be tested in other tissues and organs.

Effects of Zoledronate on Local and Systemic Production of IL-1ß, IL-18, and TNF-α in Mice and Augmentation by Lipopolysaccharide.

Bisphosphonates (BPs) containing nitrogen (N-BPs) exhibit far stronger anti-bone-resorptive effects than non-N-BPs. However, repeated administration of N-BPs causes osteonecrosis selectively in jawbones. As BPs accumulate in large amounts within inflamed bones, any N-BP released from the pool accumulated within jawbones might directly act on cells in the surrounding soft-tissues and induce inflammation or necrosis. Here, we examined the local and systemic effects of zoledronate (the most potent N-BP with the highest incidence of jawbone-necrosis) on inflammatory cytokines in mice. Locally within ear-pinnas: (i) zoledronate induced long-lasting accumulation of interleuikin-1ß (IL-1ß) and IL-18, but not tumor necrosis factor-α (TNF-α), (ii) zoledronate and lipopolysaccharide (LPS, a cell-wall component of Gram-negative bacteria) mutually augmented the productions of IL-1ß, IL-18, and TNF-α, and (iii) oxidronate (a toxic non-N-BP) by itself produced not only IL-1ß and IL-18, but also TNF-α. In systemic experiments using intraperitoneal injection of zoledronate and/or LPS, (i) zoledronate by itself increased none of the above cytokines in serum, and (ii) in mice pretreated (3 d before) with zoledronate, the LPS-induced increases in serum IL-1ß and IL-18 were greatly augmented with a delayed slight TNF-α augmentation. These results, together with previous ones, suggest that (a) pro-IL-1ß and pro-IL-18 accumulate within cells in soft-tissues exposed to N-BPs, and infection may augment not only their production, but also the release of their mature forms, (b) IL-1ß and IL-18 (possibly together with TNF-α) may play important roles in N-BP-induced inflammation and/or necrosis, and (c) mechanisms underlying the cytotoxic effects of BPs may differ between N-BPs and non-N-BPs.

The Role of Hyaluronidase for the Skin Necrosis Caused by Hyaluronic Acid Injection-Induced Embolism: A Rabbit Auricular Model Study.

BACKGROUND: Skin necrosis is considered the most serious complication of hyaluronic acid dermal filler injection procedures. To effectively treat skin necrosis, hyaluronidase injection is one of the essential preventative treatments, and yet optimal complication management remains an unmet need. Therefore, this paper investigates the effects of hyaluronidase injection timing on the treatment of skin necrosis. METHODS: In an in vitro experiment, the carbazole method was used to determine the degradation time of hyaluronic acid gels in a large volume of hyaluronidase. In vivo experimental rabbit ear models were developed to simulate the skin necrosis caused by hyaluronic acid and the test animals distributed into five groups. Except one control group, the other four groups were injected with a large volume of hyaluronidase as treatment at 2 h, 4 h, 8 h and 16 h, respectively, after models were built. The necrosis degree of models was analyzed with necrotic area and histologic examination on the postoperative 7th day. Besides, temperatures of rabbit ears were observed to demonstrate the healing process of flap models. RESULTS: The average necrotic area of flaps in the 2-h and 4-h injection groups showed a significant difference compared with that of the control group (p < 0.05; p < 0.05). The histologic examination showed that there were HA embolisms, vascular thrombolytic recanalization and arteriovenous thromboses in the survival area. In addition, the mean temperatures of the rabbit ear flaps fluctuated over time and showed clear differences between distal and proximal parts. CONCLUSIONS: The area of flap necrosis positively correlates with injection timing of the large volume of hyaluronidase. More importantly, when injection timing is within 4 h, treatment effectiveness will be significantly improved. NO LEVEL ASSIGNED: This journal requires that authors assign a level of evidence to each article. For a full description of these Evidence-Based Medicine ratings, please refer to the Table of Contents or the online Instructions to Authors www.springer.com/00266 .

Nonsurgical treatment of earlobe aging in Mowlavi stages I and II earlobe ptosis with Hyaluronic acid fillers.

BACKGROUND: Earlobe deflation caused by fat atrophy is normally treated with lipofilling, mostly in the context of facelift surgery. In this report, we aim at reporting on Hyaluronic Acid injections to treat earlobe deflation. MATERIALS AND METHODS: 16 Mowlavi Grade I and II patients were treated with HA injections, followed by molding to shape the lobule. RESULTS: Effective correction, lasting 14 months on average, is achieved. Five patients needed a touch-up procedure after 4-6 months to improve the result. CONCLUSIONS: Earlobe augmentation with HA is an ideal option for correction of earlobe atrophy in cases of Mowlavi Grades I and II ptosis. Long-lasting (about 14 months) correction is achieved with no downtime.

Evaluación de la actividad anti-inflamatoria de propóleos chileno sobre cortes histológicos de orejas de ratón / Evaluation of the anti-inflammatory activity of Chilean propolis on histological sections of mouse ears

RESUMEN: El propóleos es un producto resinoso complejo producido por las abejas Apis mellifera, el cual posee diversas actividades biológicas como inmunomodulador, antiinflamatorio, anticancerígeno, antiviral, antibacteriano, antioxidante, entre otros. El propósito del siguiente estudio fue realizar una evaluación in vivo de las propiedades antiinflamatorias de un extracto de propóleos chileno, sobre el modelo de edema auricular inducido por 13-acetato-12-O-tetradecanoilforbol (TPA) en pabellón auricular de ratón, para posterior evaluación y análisis histológico. El extracto de propóleos chileno (EEP) utilizado se obtuvo a partir de un macerado etanólico, rotaevaporado y liofilizado. Se observó que el EEP disminuyó el edema y el infiltrado inflamatorio de forma significativa. Estos resultados sugieren que el extracto etanólico de propóleos chileno posee potenciales efectos antiinflamatorios o moduladores del sistema inmunológico en edema auricular.

SUMMARY: Propolis is a complex resinous product produced by bees Apis mellifera, which has a number of biological activities such as an immunomodulator, anti-inflammatory, anticarcinogenic, antiviral, antibacterial, antioxidant, among others. The purpose of the following study was to perform an in vivo evaluation of the anti-inflammatory properties of a Chilean propolis extract, on the model of atrial edema induced 12-O-tetradecanoyl phorbol-13- acetate (TPA) in the mouse auricular pavilion, for later evaluation and histological analysis. The Chilean propolis extract (EPP) used was obtained from an ethanolic, rotaevaporated and lyophilized macerate. It was observed that the EPP significantly decreased edema and inflammatory infiltrate. These results suggest that the ethanolic extract of Chilean propolis possesses potential anti-inflammatory or modulatory effects of the immune system in atrial edema.

Exosomes of human placenta-derived mesenchymal stem cells stimulate angiogenesis.

BACKGROUND: The therapeutic potential of mesenchymal stem cells (MSCs) may be attributed partly to humoral factors such as growth factors, cytokines, and chemokines. Human term placental tissue-derived MSCs (PlaMSCs), or conditioned medium left over from cultures of these cells, have been reported to enhance angiogenesis. Recently, the exosome, which can transport a diverse suite of macromolecules, has gained attention as a novel intercellular communication tool. However, the potential role of the exosome in PlaMSC therapeutic action is not well understood. The purpose of this study was to evaluate PlaMSC-derived exosome angiogenesis promotion in vitro and in vivo. METHODS: MSCs were isolated from human term placental tissue by enzymatic digestion. Conditioned medium was collected after 48-h incubation in serum-free medium (PlaMSC-CM). Angiogenic factors present in PlaMSC-CM were screened by a growth factor array. Exosomes were prepared by ultracentrifugation of PlaMSC-CM, and confirmed by transmission electron microscopy, dynamic light scattering, and western blot analyses. The proangiogenic activity of PlaMSC-derived exosomes (PlaMSC-exo) was assessed using an endothelial tube formation assay, a cell migration assay, and reverse transcription-PCR analysis. The in-vivo angiogenic activity of PlaMSC-exo was evaluated using a murine auricle ischemic injury model. RESULTS: PlaMSC-CM contained both angiogenic and angiostatic factors, which enhanced endothelial tube formation. PlaMSC-exo were incorporated into endothelial cells; these exosomes stimulated both endothelial tube formation and migration, and enhanced angiogenesis-related gene expression. Laser Doppler blood flow analysis showed that PlaMSC-exo infusion also enhanced angiogenesis in an in-vivo murine auricle ischemic injury model. CONCLUSIONS: PlaMSC-exo enhanced angiogenesis in vitro and in vivo, suggesting that exosomes play a role in the proangiogenic activity of PlaMSCs. PlaMSC-exo may be a novel therapeutic approach for treating ischemic diseases.

Clinical Application of Earlobe Augmentation with Hyaluronic Acid Filler in the Chinese Population.

BACKGROUND: Larger earlobes, which are a symbol of "richness" in traditional Chinese culture, are favored by Chinese patients. The objective of this paper is to investigate the application of earlobe augmentation with hyaluronic acid (HA) filler injection and its clinical effects in the Chinese population. METHODS: A total of 19 patients (38 ears) who received earlobe augmentation with HA filler injections between March 2013 and March 2015 were included. The clinical effects, duration, and complications of these cases were investigated. RESULTS: All patients who received earlobe HA injections showed immediate postoperative effects with obvious morphological improvement of their earlobes. The volume of HA filler injected into each ear was 0.3-0.5 ml. The duration of the effect was 6-9 months. Two of the 19 cases (3 ears) demonstrated mild bruising at the injection site, but the bruising completely disappeared within 7 days after the injection. No vascular embolism, infection, nodule, or granuloma complications were observed in the studied group. CONCLUSION: The application of earlobe augmentation with HA filler injection is a safe, effective, simple procedure for earlobe shaping. It has an easy clinical application with good clinical prospects. LEVEL OF EVIDENCE V: This journal requires that authors assign a level of evidence to each article. For a full description of these Evidence-Based Medicine ratings, please refer to the Table of Contents or the online Instructions to Authors www.springer.com/00266 .

A quantitative model for dermal infection and oedema in BALB/c mice pinna.

BACKGROUND: Pharmaceutical industry demands innovation for developing new molecules to improve effectiveness and safety of therapeutic medicines. Preclinical assays are the first tests performed to evaluate new therapeutic molecules using animal models. Currently, there are several models for evaluation of treatments, for dermal oedema or infection. However, the most common or usual way is to induce the inflammation with chemical substances instead of infectious agents. On the other hand, this kind of models require the implementation of histological techniques and the interpretation of pathologies to verify the effectiveness of the therapy under assessment. This work was focused on developing a quantitative model of infection and oedema in mouse pinna. The infection was achieved with a strain of Streptococcus pyogenes that was inoculated in an injury induced at the auricle of BALB/c mice, the induced oedema was recorded by measuring the ear thickness with a digital micrometer and histopathological analysis was performed to verify the damage. The presence of S. pyogenes at the infection site was determined every day by culture. RESULTS: Our results showed that S. pyogenes can infect the mouse pinna and that it can be recovered at least for up to 4 days from the infected site; we also found that S. pyogenes can induce a bigger oedema than the PBS-treated control for at least 7 days; our results were validated with an antibacterial and anti-inflammatory formulation made with ciprofloxacin and hydrocortisone. CONCLUSIONS: The model we developed led us to emulate a dermal infection and allowed us to objectively evaluate the increase or decrease of the oedema by measuring the thickness of the ear pinna, and to determine the presence of the pathogen in the infection site. We consider that the model could be useful for assessment of new anti-inflammatory or antibacterial therapies for dermal infections.

Keloidectomy with core fillet flap and intralesional verapamil injection for recurrent earlobe keloids.

BACKGROUND: Earlobe keloids are usually recalcitrant to treatment and have a high rate of recurrence. Verapamil is a calcium channel antagonist that has been shown to inhibit the synthesis/secretion of extracellular matrix molecules and increase collagenase. OBJECTIVES: This prospective study was designed to evaluate the results of treatment of recurrent earlobe keloids using keloidectomy with core fillet flap and intralesional verapamil injection. METHODS: Nineteen keloids in 16 patients were treated using this technique with intralesional verapamil injection given intraoperatively, then every 2 weeks for 3 months, with postoperative follow-up for 18 months. RESULTS: Fourteen patients completed the study. Ten patients (71.4%) showed response to treatment. Four (28.6%) cases showed recurrence, two (14.2%) at the wound bed and another two (14. 2%) at the incision line. Eighty percent of responders were highly satisfied with their treatment. CONCLUSION: Keloidectomy with core fillet flap and intralesional verapamil injection is a reliable and cost-effective method in the treatment of recurrent earlobe keloids with a low rate of recurrence and high patient satisfaction.

Use of an in vitro model in tissue engineering to study wound repair and differentiation of blastema tissue from rabbit pinna.

Rabbit ear wound repair is an accepted model for studies of tissue regeneration, leading to scar less wound repair. It is believed that a specific tissue, blastema, is responsible for such interesting capacity of tissue regeneration. To test this idea further and to elucidate the cellular events happening during the ear wound repair, we designed some controlled experiments in vitro. Small pieces of the ear were punched and washed immediately with normal saline. The tissues were then cultured in the Dulbecco's Modified Eagle(')s Medium, supplemented with fetal bovine serum in control group. As a treatment vitamin A and C was used to evaluate the differentiation potency of the tissue. These tissues were fixed, sectioned, stained, and microscopically studied. Micrographs of electron microscopy provided evidences revealing dedifferentiation of certain cells inside the punched tissues after incubation in tissue culture medium. The histological studies revealed that cells of the tissue (i) can undergo cellular proliferation, (ii) differentiate to epithelial, condrogenic, and osteogenic tissues, and (iii) regenerate the wounds. These results could be used for interpretation of the possible events happening during tissue engineering and wound repair in vitro. An important goal of this study is to create a tissue engineering and tissue banking model, so that in the future it could be used in further blastema tissue studies at different levels.

The Usefulness of 0.2% Topical Nitroglycerin for Chondrodermatitis Nodularis Helicis.

BACKGROUND AND OBJECTIVE: Chondrodermatitis nodularis helicis (CNH) is a painful idiopathic degenerative condition involving the skin and cartilage of the helix or antihelix of the ear. Topical nitroglycerin 2% is a relatively recent treatment option for CNH that has produced good results, although with adverse effects (17% of cases). The use of a lower concentration would probably achieve similar results with fewer adverse effects. The aim of this study was to evaluate the effectiveness and safety of topical nitroglycerin 0.2% in the treatment of CNH. MATERIAL AND METHODS: We performed a retrospective observational study of patients treated in 2 Spanish hospitals between 2012 and 2014. The effectiveness of treatment was determined by clinical photography and assessment of symptoms using a verbal numerical rating scale. RESULTS: Of the 29 patients treated, 93% showed clinical improvement. In the group of responders, mean treatment duration was 1.8 months and mean follow-up was 5.9 months. Overall tolerance was good in all cases. CONCLUSION: Topical nitroglycerin 0.2% is an effective and well-tolerated conservative treatment option that improves the appearance of lesions and provides symptomatic relief in the majority of patients with CNH.

Trans-pinnal movement of methimazole: an in vitro study showing that methimazole can cross from the inner to outer pinna of cats.

OBJECTIVES: The aim of the study was to determine if methimazole applied in a transdermal formulation to the internal pinna will cross to the external pinna in an in vitro Franz cell model. METHODS: The ears from six cats were harvested soon after death. Whole ears were mounted onto Franz-type diffusion cells with the stratum corneum of the inner pinnae uppermost. A commercial transdermal preparation containing methimazole (0.1 ml/10 mg) was applied to the inner pinnae. At 1, 2, 4, 6, 8, 12, 18, 24 and 30 h, a 200 µl sample of reservoir solution was removed to determine the methimazole concentration by high-performance liquid chromatography. The ears were then dissected, separating the internal pinna from the cartilage and the external pinna, before the methimazole concentration was measured at each site. The thickness of the different regions of the ear was measured on paraffin histology sections. RESULTS: Mean ± SD methimazole concentrations at 30 h for the right and left ear, respectively, were: inner ear, 1.25 ± 0.53 mg/g, 0.39 ± 0.26 mg/g; cartilage, 1.36 ± 0.47 mg/g, 0.33 ± 0.20 mg/g; and outer ear, 1.0 ± 0.32 mg/g, 0.33 ± 0.14 mg/g. There was a difference between the left and right ears (P <0.001). Minimal methimazole concentrations were detected in the receptor fluid. The mean methimazole concentration absorbed by the skin after application of 10 mg was, for the right ear, 3.65 ± 1.27 mg/g and, for the left, 1.08 ± 0.27 mg/g. There was no correlation between methimazole concentrations and thickness of each region of the ear. CONCLUSIONS AND RELEVANCE: Methimazole in a lipophilic vehicle applied to the inner pinna will penetrate to the outer pinna of cats in an in vitro model, which may have safety implications for humans associated with cats treated with transdermal methimazole. Substantial inter-individual variation was found. Further research is required in the area of transdermal penetration of drugs in cats.

Dietary fatty acid content regulates wound repair and the pathogenesis of osteoarthritis following joint injury.

OBJECTIVE: The mechanisms linking obesity and osteoarthritis (OA) are not fully understood and have been generally attributed to increased weight, rather than metabolic or inflammatory factors. Here, we examined the influence of fatty acids, adipokines, and body weight on OA following joint injury in an obese mouse model. METHODS: Mice were fed high-fat diets rich in various fatty acids (FA) including saturated FAs (SFAs), ω-6 polyunsaturated FAs (PUFAs), and ω-3 PUFAs. OA was induced by destabilising the medial meniscus. Wound healing was evaluated using an ear punch. OA, synovitis and wound healing were determined histologically, while bone changes were measured using microCT. Activity levels and serum cytokines were measured at various time-points. Multivariate models were performed to elucidate the associations of dietary, metabolic and mechanical factors with OA and wound healing. RESULTS: Using weight-matched mice and multivariate models, we found that OA was significantly associated with dietary fatty acid content and serum adipokine levels, but not with body weight. Furthermore, spontaneous activity of the mice was independent of OA development. Small amounts of ω-3 PUFAs (8% by kcal) in a high-fat diet were sufficient to mitigate injury-induced OA, decreasing leptin and resistin levels. ω-3 PUFAs significantly enhanced wound repair, SFAs or ω-6 PUFAs independently increased OA severity, heterotopic ossification and scar tissue formation. CONCLUSIONS: Our results indicate that with obesity, dietary FA content regulates wound healing and OA severity following joint injury, independent of body weight, supporting the need for further studies of dietary FA supplements as a potential therapeutic approach for OA.

Facial cutaneous necrosis associated with suspected levamisole toxicity from tainted cocaine abuse.

OBJECTIVE: This study aimed to illustrate the otorhinolaryngologic manifestations of levamisole toxicity and illuminate the features of this diagnosis. METHODS: We describe a case of a known cocaine abuser with suspected levamisole toxicity who developed cutaneous necrosis of the cheeks, earlobes, nose, upper and lower lip, and the midline hard palate. We also review the existing clinical literature about this emerging phenomenon. RESULTS: Levamisole is a common adulterant in cocaine distributed in the United States and has been reported to cause microvascular thrombosis and vasculitis with resultant skin necrosis in cocaine abusers. The distribution of skin findings characteristically involves the cheeks, earlobes, nose, lips, and hard palate and responds variably to cessation of cocaine use. In its most severe cases, immune suppression and/or surgical debridement may be required. CONCLUSION: Levamisole toxicity can frequently involve the ears, nose, and throat tissues. Otorhinolaryngologists should recognize these manifestations to expeditiously diagnose and manage this condition. Failure to do so promptly can lead to complications that may necessitate reconstructive or amputation surgery.

Expression, production, and renaturation of a functional single-chain variable antibody fragment (scFv) against human ICAM-1

Intercellular adhesion molecule-1 (ICAM-1) is an important factor in the progression of inflammatory responses in vivo. To develop a new anti-inflammatory drug to block the biological activity of ICAM-1, we produced a monoclonal antibody (Ka=4.19×10−8 M) against human ICAM-1. The anti-ICAM-1 single-chain variable antibody fragment (scFv) was expressed at a high level as inclusion bodies in Escherichia coli. We refolded the scFv (Ka=2.35×10−7 M) by ion-exchange chromatography, dialysis, and dilution. The results showed that column chromatography refolding by high-performance Q Sepharose had remarkable advantages over conventional dilution and dialysis methods. Furthermore, the anti-ICAM-1 scFv yield of about 60 mg/L was higher with this method. The purity of the final product was greater than 90%, as shown by denaturing gel electrophoresis. Enzyme-linked immunosorbent assay, cell culture, and animal experiments were used to assess the immunological properties and biological activities of the renatured scFv.