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Introduction: The search for fast and efficient treatment for dermonecrotic lesions caused by the venom of the spider from the Loxosceles simillis, is a demand in health. Prednisolone is one of the most used drugs, however it has side effects. In this context, addictionally gold nanoparticles (GNPs) have anti-inflammatory, antioxidant, and antibacterial properties. The use of photobiomodulation has show to be efficient in the process of tissue repair. Therefore, the purpose of this study was to investigate the anti-inflammatory effect of photobiomodulation and GNPs associated or not with a low concentration of prednisolone in animal models of dermonecrotic lesion.Methodology: For this, rabbits with venon-induced dermonecrotic lesion were subjected to topical treatment with prednisolone + laser or GNPs + laser or Pred-GNPs + laser. The area of edema, necrosis and erythema were measured. On the last day of treatment, the animals were euthanized to remove the organs for histopathological and biochemical analysis.Results: All treatments combinations were effective in promoting the reduction of necrotic tissue and erythema.Conclusion: With this results, we suggest that the use of laser and nanoparticles, associated or not with prednisolone, should be considered for the treatment of dermonecrotic injury.

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Sphingomyelinase D (SMase D), the main toxic component of Loxosceles venom, has a well-documented role on dermonecrotic lesion triggered by envenomation with these species; however, the intracellular mechanisms involved in this event are still poorly known. Through differential transcriptomics of human keratinocytes treated with L. laeta or L. intermedia SMases D, we identified 323 DEGs, common to both treatments, as well as upregulation of molecules involved in the IL-1 and ErbB signaling. Since these pathways are related to inflammation and wound healing, respectively, we investigated the relative expression of some molecules related to these pathways by RT-qPCR and observed different expression profiles over time. Although, after 24 h of treatment, both SMases D induced similar modulation of these pathways in keratinocytes, L. intermedia SMase D induced earlier modulation compared to L. laeta SMase D treatment. Positive expression correlations of the molecules involved in the IL-1 signaling were also observed after SMases D treatment, confirming their inflammatory action. In addition, we detected higher relative expression of the inhibitor of the ErbB signaling pathway, ERRFI1, and positive correlations between this molecule and pro-inflammatory mediators after SMases D treatment. Thus, herein, we describe the cell pathways related to the exacerbation of inflammation and to the failure of the wound healing, highlighting the contribution of the IL-1 signaling pathway and the ERRFI1 for the development of cutaneous loxoscelism.

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Sphingomyelinase D (SMase D), the main toxic component of Loxosceles venom, has a well-documented role on dermonecrotic lesion triggered by envenomation with these species; however, the intracellular mechanisms involved in this event are still poorly known. Through differential transcriptomics of human keratinocytes treated with L. laeta or L. intermedia SMases D, we identified 323 DEGs, common to both treatments, as well as upregulation of molecules involved in the IL-1 and ErbB signaling. Since these pathways are related to inflammation and wound healing, respectively, we investigated the relative expression of some molecules related to these pathways by RT-qPCR and observed different expression profiles over time. Although, after 24 h of treatment, both SMases D induced similar modulation of these pathways in keratinocytes, L. intermedia SMase D induced earlier modulation compared to L. laeta SMase D treatment. Positive expression correlations of the molecules involved in the IL-1 signaling were also observed after SMases D treatment, confirming their inflammatory action. In addition, we detected higher relative expression of the inhibitor of the ErbB signaling pathway, ERRFI1, and positive correlations between this molecule and pro-inflammatory mediators after SMases D treatment. Thus, herein, we describe the cell pathways related to the exacerbation of inflammation and to the failure of the wound healing, highlighting the contribution of the IL-1 signaling pathway and the ERRFI1 for the development of cutaneous loxoscelism.

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Resumen ANTECEDENTES: el loxoscelismo es una intoxicación por la mordedura de la araña Loxosceles reclusa, cuyo veneno contiene esfingomielinasa-D, causante de hemólisis y necrosis. Se reporta una serie de casos que describen su evolución clínica y respuesta al tratamiento. OBJETIVO: describir la evolución y características clínicas de pacientes con loxoscelismo sistémico y dermonecrótico, su respuesta al tratamiento y las complicaciones. PACIENTES Y MÉTODO: estudio que incluyó el análisis descriptivo de pacientes tratados en el servicio de Medicina Interna, Unidad Médica de Alta Especialidad Dr. Antonio Fraga Mouret, Centro Médico Nacional La Raza, de 2010 a 2015. RESULTADOS: se atendieron ocho hombres (67%) y cuatro mujeres (33%), con edad media de 39.4 años (16-83 años). El sitio de mordedura en cinco casos (42%) fue el miembro pélvico izquierdo; cuatro casos en el miembro torácico derecho (33%), dos en la cara (17%) y uno en el miembro torácico izquierdo (9%). Nueve casos provenían del Estado de México, dos de la Ciudad de México y uno de Hidalgo. Manifestación clínica: flictenas (cinco casos), edema y eritema (tres), placa liveloide (tres) y necrosis (un caso). Nueve (75%) pacientes se trataron inicialmente en la unidad de cuidados intensivos. Se reportaron las siguientes complicaciones sistémicas: renales (67%), pulmonares con administración de aminas (33%) y hematológicas (8%). Diez casos recibieron faboterápico, con media de 1.5 viales (0 a 4); cinco casos (42%) recibieron dapsona y cuatro de ellos (33%) padecieron metahemoglobinemia; once (92%) pacientes requirieron lavado-desbridación y 7 (58%) injerto cutáneo; cuatro pacientes (33%) tuvieron infección agregada de la herida. El promedio de estancia hospitalaria fue 16.2 días (3 a 40 días). CONCLUSIÓN: la evolución y pronóstico de esta afección depende de una sospecha inicial, diagnóstico y tratamiento oportunos. El loxoscelismo debe incluirse en los diagnósticos diferenciales de lesiones necróticas y progresivas, con o sin afección sistémica.

Abstract BACKGROUND: Loxoscelism is a poisoning caused by the bite of Loxosceles recluse spider, whose venom contains sphingomyelinaseD, causing hemolysis and necrosis. We report a case series describing their clinical course and response to treatment. OBJECTIVE: To describe the evolution and clinical characteristics of patients with systemic and dermonecrotic loxoscelism, their response to treatment and complications. PATIENTS AND METHOD: A descriptive analysis of patients treated in the Internal Medicine Service, Centro Médico Nacional La Raza, from 2010 to 2015. RESULTS: A total of 8 men (67%) and 4 women (33%) were included. Mean age was 39.4 years (16-83 years). Bite site was left pelvic limb in 5 cases (42%), 4 in the right forelimb (33%), 2 in the face (17%) and 1 in left forelimb (9%). Nine cases came from Estado de México, 2 from Mexico City and 1 from Hidalgo. Initial manifestations included blisters (five cases), edema and erythema (three cases), liveloide plate (three cases) and necrosis (one case). Nine (75%) patients were initially managed in ICU. Systemic complications were renal (67%), lung with use of amines (33%) and hematological (8%). Ten cases were treated with fabotherapy, with an average of 1.5 vials (0-4). Five cases (42%) received dapsone and 4 of them (33%) developed methemoglobinemia. Eleven (92%) required surgical washing and debridement and 7 (58%) skin graft; four patients (33%) had secondary wound infection. Average hospital stay was 16.2 days (3-40 days). CONCLUSIONS: The evolution and prognosis depends on initial suspicion early diagnosis and treatment. Loxoscelism should be included in the differential diagnosis of progressive necrotic lesions, with or without systemic involvement.

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UNLABELLED: Brown spider (Loxosceles genus) bites have been reported worldwide. The venom contains a complex composition of several toxins, including phospholipases-D. Native or recombinant phospholipase-D toxins induce cutaneous and systemic loxoscelism, particularly necrotic lesions, inflammatory response, renal failure, and hematological disturbances. Herein, we describe the cloning, heterologous expression and purification of a novel phospholipase-D toxin, LiRecDT7 in reference to six other previously described in phospholipase-D toxin family. The complete cDNA sequence of this novel brown spider phospholipase-D isoform was obtained and the calculated molecular mass of the predicted mature protein is 34.4 kDa. Similarity analyses revealed that LiRecDT7 is homologous to the other dermonecrotic toxin family members particularly to LiRecDT6, sharing 71% sequence identity. LiRecDT7 possesses the conserved amino acid residues involved in catalysis except for a conservative mutation (D233E) in the catalytic site. Purified LiRecDT7 was detected as a soluble 36 kDa protein using anti-whole venom and anti-LiRecDT1 sera, indicating immunological cross-reactivity and evidencing sequence-epitopes identities similar to those of other phospholipase-D family members. Also, LiRecDT7 exhibits sphingomyelinase activity in a concentration dependent-manner and induces experimental skin lesions with swelling, erythema and dermonecrosis. In addition, LiRecDT7 induced a massive inflammatory response in rabbit skin dermis, which is a hallmark of brown spider venom phospholipase-D toxins. Moreover, LiRecDT7 induced in vitro hemolysis in human erythrocytes and increased blood vessel permeability. These features suggest that this novel member of the brown spider venom phospholipase-D family, which naturally contains a mutation (D233E) in the catalytic site, could be useful for future structural and functional studies concerning loxoscelism and lipid biochemistry. HIGHLIGHTS: 1- Novel brown spider phospholipase-D recombinant toxin contains a conservative mutation (D233E) on the catalytic site. 2-LiRecDT7 shares high identity level with isoforms of Loxosceles genus. 3-LiRecDT7 is a recombinant protein immunodetected by specific antibodies to native and recombinant phospholipase-D toxins. 4-LiRecDT7 shows sphingomyelinase-D activity in a concentration-dependent manner, but less intense than other isoforms. 5-LiRecDT7 induces dermonecrosis and inflammatory response in rabbit skin. 6-LiRecDT7 increases vascular permeability in mice. 7-LiRecDT7 triggers direct complement-independent hemolysis in erythrocytes.

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Envenomations caused by Loxosceles (brown spider) have been reported throughout the world. Clinical signs associated to bites of these spiders involve dermonecrotic lesions and intense local inflammatory response, besides systemic manifestations such as intravascular hemolysis, thrombocytopenia, disseminated intravascular coagulation and acute renal failure. The present study aimed to report and to describe dermonecrotic lesions probably caused by a Loxosceles envenomation in a four year-old poodle female dog, treated at the Dermatology Service of the Veterinary Hospital of the Veterinary Medicine and Animal Husbandry School, São Paulo State University, Botucatu, Brazil. Initially, the animal presented two skin lesions with blackish aspect that evolved into ulcerative crusts. The owner reported the presence of a brown spider near the place where the animal spent most of the time. Histological examination of lesions revealed necrosis of the epidermis extending to adnexa and panniculi, which is compatible with Loxosceles bite reaction. The animal was treated with systemic antibiotic and local curatives. Lesions healed by second intention in two months.

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Envenomations caused by Loxosceles (brown spider) have been reported throughout the world. Clinical signs associated to bites of these spiders involve dermonecrotic lesions and intense local inflammatory response, besides systemic manifestations such as intravascular hemolysis, thrombocytopenia, disseminated intravascular coagulation and acute renal failure. The present study aimed to report and to describe dermonecrotic lesions probably caused by a Loxosceles envenomation in a four year-old poodle female dog, treated at the Dermatology Service of the Veterinary Hospital of the Veterinary Medicine and Animal Husbandry School, São Paulo State University, Botucatu, Brazil. Initially, the animal presented two skin lesions with blackish aspect that evolved into ulcerative crusts. The owner reported the presence of a brown spider near the place where the animal spent most of the time. Histological examination of lesions revealed necrosis of the epidermis extending to adnexa and panniculi, which is compatible with Loxosceles bite reaction. The animal was treated with systemic antibiotic and local curatives. Lesions healed by second intention in two months.(AU)

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