Licorice zinc suppresses melanogenesis via inhibiting the activation of P38MAPK and JNK signaling pathway in Balb/c mice skin

Purpose: The active melanocytes in the skin were affected by hormones and ultraviolet (UV) irradiation. Licorice zinc has a whitening effect, which may have a prominent potential in the treatment of pigmented skin disease. Methods: Modeling chloasma C57BL/6J mice by daily progesterone injection (15 mg/kg) and ultraviolet B (UVB) irradiation (λ = 312 nm, 2 h/day) for 30 days. Then, mice were given 0.65, 1.3, and 2.6 (g/kg) of licorice zinc and tranexamic acid 250 mg daily by oral administration for 14 days, respectively. Hematoxylin and eosin and Fontana-Masson staining, and Western blotting (WB) were performed to test the inhibitory of melanogenesis and activation of c-Jun-N-terminal (JNK)/p38 mitogen-activated protein kinases (MAPK) for licorice zinc. Melanogenesis was induced by α-melanocyte-stimulating hormone in vitro. Cell counting kit-8, melanin content determination, and WB were performed to verify the inhibitory effect of licorice zinc on melanogenesis. Results: The present study showed that licorice zinc decreased melanin formation, cutaneous tissue injury, and the phosphorylation of JNK and P38MAPK, which was caused by UVB irradiation in vivo. In vitro, licorice zinc showed opposite effects from JNK/p38 activator. Meanwhile, tyrosinase-related protein-1, tyrosinase, and microphthalmia-associated transcription factor were decreased too. Conclusions: Licorice zinc induced a decrease in melanin synthesis by inhibiting the JNK and the P38MAPK signaling pathway, suggesting licorice zinc is a potential agent of anti-chloasma.

Transient skin and fur color change after trichotomy of the sacrococcygeal region for epidural anesthesia in a domestic cat (Felis catus) / Mudança transitória da cor da pele e pelo após tricotomia da região sacrococcígea para anestesia epidural em gata doméstica (Felis catus)

A clinical case in which skin and hair color change occurred after sacrococcygeal epidural anesthesia in a nine-month-old Siamese cross queen undergoing ovariohysterectomy (OHE) is described. Six weeks after surgery, during a re-check, it was noted that in the sacrococcygeal region the color of the skin and new hair growth was dark with a color comparable to the color present on the body extremities (muzzle, pinnae, legs, and tail). The skin and new hair growth of the shaved abdomen presented a standard color. The key enzyme of the melanogenic pathway in mammals is tyrosinase (TYR), and the Siamese temperature-sensitive phenotype is the result of genetic mutations that makes TYR function thermolabile. The activity of TYR in these cats is limited to the extremities where the temperature is lower while pigment production is impaired in the other body areas. The trichotomy of the sacrococcygeal region performed during wintertime in an outdoor cat was probably the trigger for increased activity of TYR in this area promoting pigment production. The absence of the same alterations in the abdominal area may be justified by less exposure of that region to the external environment, as well as to the feline habits of sedentarism, that avoid significant cooling in these regions. This report highlights the importance of taking this type of occurrence into account when performing an epidural in the Siamese cat breed. Also, to avoid skin color change in this breed, the authors recommend a midline abdominal instead of a flank approach to perform OHE.(AU)

Descreve-se um caso clínico no qual ocorreu mudança na cor da pele e do pelo após anestesia epidural sacrococcígea numa gata cruzada de raça Siamês de nove meses submetida à ovariohisterectomia (OVH). Seis semanas após a cirurgia, durante uma avaliação pós-operatória, notou-se que na região sacrococcígea, a cor da pele e o crescimento do pelo apresentavam uma cor escura, comparável à das extremidades do corpo (face, orelhas, membros e cauda). A pele e o crescimento do pelo do abdómen, que também havia sido tosquiado, apresentavam uma cor padrão. A enzima chave da via melanogênica em mamíferos é a tirosinase (TYR) e o fenótipo siamês sensível à temperatura é o resultado de mutações genéticas que tornam a função TYR termolábil. A atividade da TYR nestes gatos é limitada às extremidades onde a temperatura é mais baixa, enquanto a produção de pigmento é prejudicada em outras áreas do corpo. A tricotomia da região sacrococcígea realizada durante o inverno nesta gata com acesso livre ao ambiente externo, provavelmente determinou um aumento da atividade da TYR nesta área, promovendo a produção de pigmento. A ausência das mesmas alterações na região abdominal pode ser justificada pela menor exposição desta área do corpo ao ambiente externo, também devido aos hábitos felinos de sedentarismo, que evitam resfriamento significativo nestas regiões. Este relato destaca a importância de se levar em consideração a possibilidade deste tipo de ocorrência quando da realização de uma anestesia epidural nesta raça de gatos. Além disso, os autores recomendam uma abordagem abdominal na linha média ao invés de uma abordagem de flanco para realizar a OVH nesta raça, a fim de evitar a alteração da cor da pele.(AU)

Transient skin and fur color change after trichotomy of the sacrococcygeal region for epidural anesthesia in a domestic cat (Felis catus) / Mudança transitória da cor da pele e pelo após tricotomia da região sacrococcígea para anestesia epidural em gata doméstica (Felis catus)

A clinical case in which skin and hair color change occurred after sacrococcygeal epidural anesthesia in a nine-month-old Siamese cross queen undergoing ovariohysterectomy (OHE) is described. Six weeks after surgery, during a re-check, it was noted that in the sacrococcygeal region the color of the skin and new hair growth was dark with a color comparable to the color present on the body extremities (muzzle, pinnae, legs, and tail). The skin and new hair growth of the shaved abdomen presented a standard color. The key enzyme of the melanogenic pathway in mammals is tyrosinase (TYR), and the Siamese temperature-sensitive phenotype is the result of genetic mutations that makes TYR function thermolabile. The activity of TYR in these cats is limited to the extremities where the temperature is lower while pigment production is impaired in the other body areas. The trichotomy of the sacrococcygeal region performed during wintertime in an outdoor cat was probably the trigger for increased activity of TYR in this area promoting pigment production. The absence of the same alterations in the abdominal area may be justified by less exposure of that region to the external environment, as well as to the feline habits of sedentarism, that avoid significant cooling in these regions. This report highlights the importance of taking this type of occurrence into account when performing an epidural in the Siamese cat breed. Also, to avoid skin color change in this breed, the authors recommend a midline abdominal instead of a flank approach to perform OHE.(AU)

Descreve-se um caso clínico no qual ocorreu mudança na cor da pele e do pelo após anestesia epidural sacrococcígea numa gata cruzada de raça Siamês de nove meses submetida à ovariohisterectomia (OVH). Seis semanas após a cirurgia, durante uma avaliação pós-operatória, notou-se que na região sacrococcígea, a cor da pele e o crescimento do pelo apresentavam uma cor escura, comparável à das extremidades do corpo (face, orelhas, membros e cauda). A pele e o crescimento do pelo do abdómen, que também havia sido tosquiado, apresentavam uma cor padrão. A enzima chave da via melanogênica em mamíferos é a tirosinase (TYR) e o fenótipo siamês sensível à temperatura é o resultado de mutações genéticas que tornam a função TYR termolábil. A atividade da TYR nestes gatos é limitada às extremidades onde a temperatura é mais baixa, enquanto a produção de pigmento é prejudicada em outras áreas do corpo. A tricotomia da região sacrococcígea realizada durante o inverno nesta gata com acesso livre ao ambiente externo, provavelmente determinou um aumento da atividade da TYR nesta área, promovendo a produção de pigmento. A ausência das mesmas alterações na região abdominal pode ser justificada pela menor exposição desta área do corpo ao ambiente externo, também devido aos hábitos felinos de sedentarismo, que evitam resfriamento significativo nestas regiões. Este relato destaca a importância de se levar em consideração a possibilidade deste tipo de ocorrência quando da realização de uma anestesia epidural nesta raça de gatos. Além disso, os autores recomendam uma abordagem abdominal na linha média ao invés de uma abordagem de flanco para realizar a OVH nesta raça, a fim de evitar a alteração da cor da pele.(AU)

A fatal pneumonia due to coinfection of Pseudomonas putida and Staphylococcus pseudintermedius in a laboratory beagle dog

Background: Pseudomonas putida (P. putida) is widely distributed in the environment, and sometimes caused nosocomialinfections in human beings, but no case of infection has been reported in beagle dogs. Staphylococcus pseudintermedius(S. pseudintermedius) is a natural cutaneous bacterium in dogs and occasionally causes purulent infections of the skin yetrarely causes pneumonia. Both bacteria are opportunistic pathogens. Dogs, even well-controlled laboratory beagle dogs,maybe infected by the bacterium in certain conditions like this report. In order to provide information and give suggestionto veterinarians involved in dogs study, a complete profile of the coinfection was drawn in this report.Case: It is presented a case of an 8-month-old beagle dog, weighing 6 kg that suffered from coinfection of P. putida andS. pseudintermedius during a treatment of chemotherapy. The animal was confirmed as normal by appearance, physicalexamination and laboratory tests before arrival according to the applicable guidelines. After 14-day acclimation period, theanimal was administrated with a tyrosinase inhibitor once daily via oral gavage. From Day 8, coughing, decreased activity, hyporeflexia, squinting, shortness of breath (abdominal breathing), and discharge around the nose as well as cracklesin the lung and rapid heart rate were noted. Since the poor conditions progressed quickly and have not been improved bytreatment of ceftriaxone and dexamethasone. On Day 9, the animal was euthanized for humanitarian reasons. To define thepathogen, hilar lymph node and thoracic swab were collected for bacteria isolation and purification in special mediums,and at last characterized by Gram staining and 16s rRNA gene sequence analysis and positive PCR-restriction fragmentlength polymorphism. In clinical pathological examination, an increase in WBC, neutrophils, lymphocytes, monocytes...

A fatal pneumonia due to coinfection of Pseudomonas putida and Staphylococcus pseudintermedius in a laboratory beagle dog

Background: Pseudomonas putida (P. putida) is widely distributed in the environment, and sometimes caused nosocomialinfections in human beings, but no case of infection has been reported in beagle dogs. Staphylococcus pseudintermedius(S. pseudintermedius) is a natural cutaneous bacterium in dogs and occasionally causes purulent infections of the skin yetrarely causes pneumonia. Both bacteria are opportunistic pathogens. Dogs, even well-controlled laboratory beagle dogs,maybe infected by the bacterium in certain conditions like this report. In order to provide information and give suggestionto veterinarians involved in dogs study, a complete profile of the coinfection was drawn in this report.Case: It is presented a case of an 8-month-old beagle dog, weighing 6 kg that suffered from coinfection of P. putida andS. pseudintermedius during a treatment of chemotherapy. The animal was confirmed as normal by appearance, physicalexamination and laboratory tests before arrival according to the applicable guidelines. After 14-day acclimation period, theanimal was administrated with a tyrosinase inhibitor once daily via oral gavage. From Day 8, coughing, decreased activity, hyporeflexia, squinting, shortness of breath (abdominal breathing), and discharge around the nose as well as cracklesin the lung and rapid heart rate were noted. Since the poor conditions progressed quickly and have not been improved bytreatment of ceftriaxone and dexamethasone. On Day 9, the animal was euthanized for humanitarian reasons. To define thepathogen, hilar lymph node and thoracic swab were collected for bacteria isolation and purification in special mediums,and at last characterized by Gram staining and 16s rRNA gene sequence analysis and positive PCR-restriction fragmentlength polymorphism. In clinical pathological examination, an increase in WBC, neutrophils, lymphocytes, monocytes...(AU)

Leucismo em jacaretinga: relato de caso / Leucism in jacaretinga: case report

Dentre os crocodilianos com ocorrência no Brasil, o Caiman crocodilus (Linnaeus, 1758) é a espécie de maior frequência, porém sua presença em estudos científicos restringe-se a levantamentos faunísticos, na maioria das vezes. O leucismo, também conhecido como albinismo parcial, é uma anomalia cromática ainda pouco conhecida. Indivíduos portadores dos genes que condicionam a doença apresentam ausência de pigmentação em uma parte ou em todo o corpo, porém os olhos não apresentam alteração. O objetivo deste trabalho foi realizar o primeiro registro de Caiman crocodilus com leucismo no mundo.(AU)

Among crocodilians in Brazil, Caiman crocodilus (Linnaeus, 1758) is the most frequent species, but its presence in scientific studies is mostly restricted to faunal surveys. Leukism, also known as partial albinism, is a still little known chromatic anomaly. Individuals with genes that condition the disease present no pigmentation in one part or in the whole body, but the eyes did not change. The objective of this work was to perform the first record of Caiman crocodilus with leucismo in the world.(AU)

Leucismo em jacaretinga: relato de caso / Leucism in jacaretinga: case report

Dentre os crocodilianos com ocorrência no Brasil, o Caiman crocodilus (Linnaeus, 1758) é a espécie de maior frequência, porém sua presença em estudos científicos restringe-se a levantamentos faunísticos, na maioria das vezes. O leucismo, também conhecido como albinismo parcial, é uma anomalia cromática ainda pouco conhecida. Indivíduos portadores dos genes que condicionam a doença apresentam ausência de pigmentação em uma parte ou em todo o corpo, porém os olhos não apresentam alteração. O objetivo deste trabalho foi realizar o primeiro registro de Caiman crocodilus com leucismo no mundo.(AU)

Among crocodilians in Brazil, Caiman crocodilus (Linnaeus, 1758) is the most frequent species, but its presence in scientific studies is mostly restricted to faunal surveys. Leukism, also known as partial albinism, is a still little known chromatic anomaly. Individuals with genes that condition the disease present no pigmentation in one part or in the whole body, but the eyes did not change. The objective of this work was to perform the first record of Caiman crocodilus with leucismo in the world.(AU)

Molecular detection of albinism gene in Brazilian buffalo herds (Bubalus bubalis) / Detecção molecular do gene do albinismo em rebanhos de búfalos (Bubalus bubalis) do Brasil

Albinism is a genetic disease characterized by deficient melanin production making affected animals more susceptible to skin problems, negatively influencing production systems of the same. In buffalo, a nonsense mutation (c.1431G>A) in the tyrosinase gene was already described, which is responsible for the oculocutaneous albinism buffalo phenotype. However, prevalence studies have never been performed for this anomaly in Brazil. Therefore, the objective of this study was to investigate this mutation in buffalo herd in Brazil. Of the 315 buffalo tested with no albinism phenotype evident, 11 (3.5%) were heterozygous for the mutation and none were mutated homozygous, showing the existence of the albinism gene in buffalo production herds and proving the importance of prevalence studies for hereditary diseases in order to prevent the dissemination of these same genes and their negative productivity consequences.(AU)

O Albinismo é uma doença genética caracterizada pela deficiência na produção de melanina, o que torna os animais afetados mais susceptíveis a problemas cutâneos e influencia negativamente a criação destes animais. A mutação nonsense (c.1431G>A) no gene da tyrosinase já foi descrita como responsável pelo albinismo oculocutâneo em búfalos, entretanto estudos prévios sobre a prevalência dessa mutação ainda não foram realizados no Brasil. Portanto, o objetivo deste estudo foi avaliar a presença desta mutação em uma população de búfalos brasileiros. Foram genotipados 315 búfalos clinicamente normais, ou seja, sem o fenótipo albino evidente. Desses, 11 (3,5%) eram heterozigotos para a mutação (N/TYR) e os demais eram homozigotos selvagens (N/N). Este resultado demonstra que o alelo mutado para o albinismo em búfalo está presente no rebanho brasileiro e aponta a importância de estudos de prevalência de enfermidades hereditárias com o objetivo de prevenir a disseminação desses alelos mutados, minimizando os prejuízos.(AU)

Molecular detection of albinism gene in Brazilian buffalo herds (Bubalus bubalis) / Detecção molecular do gene do albinismo em rebanhos de búfalos (Bubalus bubalis) do Brasil

Albinism is a genetic disease characterized by deficient melanin production making affected animals more susceptible to skin problems, negatively influencing production systems of the same. In buffalo, a nonsense mutation (c.1431G>A) in the tyrosinase gene was already described, which is responsible for the oculocutaneous albinism buffalo phenotype. However, prevalence studies have never been performed for this anomaly in Brazil. Therefore, the objective of this study was to investigate this mutation in buffalo herd in Brazil. Of the 315 buffalo tested with no albinism phenotype evident, 11 (3.5%) were heterozygous for the mutation and none were mutated homozygous, showing the existence of the albinism gene in buffalo production herds and proving the importance of prevalence studies for hereditary diseases in order to prevent the dissemination of these same genes and their negative productivity consequences.(AU)

O Albinismo é uma doença genética caracterizada pela deficiência na produção de melanina, o que torna os animais afetados mais susceptíveis a problemas cutâneos e influencia negativamente a criação destes animais. A mutação nonsense (c.1431G>A) no gene da tyrosinase já foi descrita como responsável pelo albinismo oculocutâneo em búfalos, entretanto estudos prévios sobre a prevalência dessa mutação ainda não foram realizados no Brasil. Portanto, o objetivo deste estudo foi avaliar a presença desta mutação em uma população de búfalos brasileiros. Foram genotipados 315 búfalos clinicamente normais, ou seja, sem o fenótipo albino evidente. Desses, 11 (3,5%) eram heterozigotos para a mutação (N/TYR) e os demais eram homozigotos selvagens (N/N). Este resultado demonstra que o alelo mutado para o albinismo em búfalo está presente no rebanho brasileiro e aponta a importância de estudos de prevalência de enfermidades hereditárias com o objetivo de prevenir a disseminação desses alelos mutados, minimizando os prejuízos.(AU)

Phenoloxidase activity and haemolymph cytology in honeybees challenged with a virus suspension (deformed wings virus DWV) or phosphate buffered suspension (PBS) / Atividade fenoloxidase e citologia de hemolinfa em abelhas infectadas com uma suspensão de vírus (DWV deformed wings) ou suspensão tamponada com fosfato (PBS)

The innate immune system of honeybees mainly consists in antimicrobial peptides, cellular immunity and melanisation. In order to investigate the immune response of honeybees to immune stressors, three stress degrees were tested. Newly emerged bees naturally DWV-infected were collected from a Varroa mite-free apiary and divided into three experimental groups: naturally DWV infected bees, PBS injected bees, and artificially DWV super infected bees. Phenoloxidase activity and haemolymph cellular subtype count were investigated. Phenoloxidase activity was highest (P<0.05) in DWV-superinfected bees, and the haemocyte population differed within the three observed groups. Although, immune responses following DWV infection have still not been completely clarified, this investigation sheds light on the relation between cell immunity and the phenoloxidase activity of DWV-naturally infected honeybees exposed to additional stress such as injury and viral superinfection.(AU)

O sistema imune inato das abelhas consiste principalmente em peptídeos antimicrobianos, imunidade celular e melanização. Para investigar a resposta imune das abelhas a estressores imunológicos, foram testados três graus de estresse. Abelhas recém-emergidas naturalmente infectadas por DWV foram coletadas de um apiário livre de Varroa e divididas em três grupos experimentais: abelhas naturalmente infectadas por DWV, abelhas injetadas com PBS e abelhas superinfectadas artificialmente com DWV. A atividade de fenoloxidase e a contagem de subtipos celulares de hemolinfa foram investigadas. A atividade da fenoloxidase foi maior (P<0,05) nas abelhas super-infectadas com DWV, e a população de hemócitos diferiu entre os três grupos observados. Embora as respostas imunes após a infecção pelo DWV ainda não tenham sido completamente esclarecidas, esta investigação lança luz sobre a relação entre a imunidade celular e a atividade da fenoloxidase das abelhas infectadas naturalmente pelo DWV, expostas a estresse adicional, como lesão e superinfecção viral.(AU)

Dalbergia ecastaphyllum leaf extracts: in vitro inhibitory potential against enzymes related to metabolic syndrome, inflammation and neurodegenerative diseases

For the first time, the anti-hemolytic activity and the enzyme inhibitory activities of Dalbergia ecastaphyllum leaves extracts were tested against α-amylase, α-glucosidase, lipase, acetylcholinesterase, butyrylcholinesterase, tyrosinase and hyaluronidase. The phenolic profile of the obtained extracts was also investigated by high-performance liquid chromatography with photodiode array detection (HPLC-PAD). The extracts showed inhibitory activity against all enzymes evaluated, with the highest inhibitory activity reported for the enzyme hyaluronidase (28.28 ± 2.43 to 72.19 ± 1.40 μg mL-1). The obtained extracts also demonstrate anti-hemolytic activity (52.22 ± 1.62 to 71.17 ± 1.82%). Among the phenolic compounds identified, protocatechuic, vanillic and β-resorcylic acids were the most abundant (1.13 ± 0.06 to 2.53 ± 0.06, 0.90 ± 0.06 to 2.19 ± 0.06 and 1.03 ± 1.62 to 22.11 ± 1.62 mg L-1, respectively). In the statistical analysis, a significant correlation was found between the flavonoids content and all enzymes inhibitory activities. The present study showed that D. ecastaphyllum leaves extracts may have the potential to be used in the therapeutic treatment of several diseases such as Alzheimer, Parkinson, type 2 diabetes mellitus, hyperglycemia, and pigmentation, as well as those associated with oxidative stress.

Canine oral osteocartilaginous malignant amelanotic melanoma with pulmonary metastasis

Background: Melanomas are typically malignant neoplasms commonly observed in the oral cavity of dogs. The classical presentation of melanomas with characteristic melanin pigmentation is easy to diagnose; however, in some cases, the lack of melanin production in the amelanotic oral tumors cause a delay in establishing the precocious diagnosis and consequent treatment. The aim of this report was to evaluate the histopathological and immunohistochemical aspects of an oral amelanotic melanoma with osteocartilaginous formation and metastasis in a dog, in a temporal way.Case: A 10-year-old male German Shepherd dog, presenting mouth bleeding with an amelanotic melanoma located between the upper incisors was received at the Veterinary Hospital of the Catholic University Dom Bosco (UCDB), Campo Grande, MS, Brazil. The animal was clinically evaluated and radiography was performed. The tumor was surgically removed and a sample was collected for histopathological examination that revealed spindloid and some epithelioid morphological cell types surrounded by a fibromatous matrix with moderate amounts of fibrovascular stroma. Approximately 1 month after surgical removal, recovery of the tumor was observed, and a second clinical analysis and collection of sample were performed. These procedures were repeated three times showing the same histopathological characteristics added by myxoid, chondroid, fibromatous tissue, and small groups of chondrocytes as well as central areas of irregular mineralized spicules. X-ray examination revealed proliferative and lytic bone infiltration in the jaw. Immunohistochemical analysis for melanocytic differentiation markers was performed showing positivity to Melan-A, tyrosinase and HMB-45 immunoreactivity, while no S100 reactivity was detected. After 11 months of the first biopsy, pleural effusion and radiopaque disseminated nodules of 1cm in the lungs were detected by X-ray. The animal died and necropsy was conducted.[...](AU)

Canine oral osteocartilaginous malignant amelanotic melanoma with pulmonary metastasis

Background: Melanomas are typically malignant neoplasms commonly observed in the oral cavity of dogs. The classical presentation of melanomas with characteristic melanin pigmentation is easy to diagnose; however, in some cases, the lack of melanin production in the amelanotic oral tumors cause a delay in establishing the precocious diagnosis and consequent treatment. The aim of this report was to evaluate the histopathological and immunohistochemical aspects of an oral amelanotic melanoma with osteocartilaginous formation and metastasis in a dog, in a temporal way.Case: A 10-year-old male German Shepherd dog, presenting mouth bleeding with an amelanotic melanoma located between the upper incisors was received at the Veterinary Hospital of the Catholic University Dom Bosco (UCDB), Campo Grande, MS, Brazil. The animal was clinically evaluated and radiography was performed. The tumor was surgically removed and a sample was collected for histopathological examination that revealed spindloid and some epithelioid morphological cell types surrounded by a fibromatous matrix with moderate amounts of fibrovascular stroma. Approximately 1 month after surgical removal, recovery of the tumor was observed, and a second clinical analysis and collection of sample were performed. These procedures were repeated three times showing the same histopathological characteristics added by myxoid, chondroid, fibromatous tissue, and small groups of chondrocytes as well as central areas of irregular mineralized spicules. X-ray examination revealed proliferative and lytic bone infiltration in the jaw. Immunohistochemical analysis for melanocytic differentiation markers was performed showing positivity to Melan-A, tyrosinase and HMB-45 immunoreactivity, while no S100 reactivity was detected. After 11 months of the first biopsy, pleural effusion and radiopaque disseminated nodules of 1cm in the lungs were detected by X-ray. The animal died and necropsy was conducted.[...]

Malignant melanoma in albino water buffalo (Bubalus bubalis) / Melanoma maligno em búfalos (Bubalus bubalis) albinos

Two albino water buffalo affected by multiple melanocytic tumors in Southern Brazil are described. Grossly, there were multiple dark tumors within the skin, skeletal muscle, lungs, and lymph nodes. The tumor was also present in the pericardial sac, renal capsule, mediastinum and pleura. Microscopically, the tumors consisted of polyhedral epithelioid cells arranged in solid nests or interwoven fascicles supported by a thin and sparse collagenous stroma. The cytoplasm was eosinophilic and sometimes contained varying amounts of melanin pigment. The mitotic rates were low. Immunohistochemistry staining with Tyrosinase, Melan-A, Vimentin, S-100 protein, and neurofilament were positive. It is possible that the polymorphisms related to pigmentation in albino buffalo contributed with a high risk of developing melanoma as suggested in humans.(AU)

Descrevem-se melanomas múltiplos em dois búfalos albinos da raça Murrah no Sul do Brasil. Macroscopicamente havia múltiplos tumores enegrecidos no músculo esquelético, pulmões, linfonodos, saco pericárdio, cápsula renal, mediastino e pleura. Microscopicamente foram observadas células epitelióides poliédricas dispostas em ninhos sólidos ou fascículos entrelaçados sustentadas por um estroma colagenoso fino e esparso. O citoplasma era eosinofílico e por vezes continham quantidades variáveis de pigmento de melanina. A taxa de mitose foi baixa. Na imuno-histoquímica houve imunomarcação positiva utilizando anticorpo anti-tirosinase, anti-Melan-A, anti-vimentina, anti-proteína S-100 e antineurofilamento. É possível que o polimorfismo relacionado com o gene da pigmentação em búfalos albinos tenha contribuído para o maior risco de desenvolvimento de melanomas, como foi sugerido em humanos.(AU)

TRIAGEM DE EXTRATOS VEGETAIS DE PLANTAS BRASILEIRAS INIBIDORES DA TIROSINASE

As desordens pigmentares são manifestações cutâneas que alteram a tonalidade da pele e resultam em desconforto estético. Substâncias com atividade inibitória no processo da melanogênese são utilizadas no tratamento das hipercromias, especialmente inibidores da enzima tirosinase. Os despigmentantes utilizados atualmente não são suficientemente eficazes e novas pesquisas em busca de princípios ativos inibidores da melanogênese mais potentes e seguros são necessárias. O presente trabalho teve como objetivo a identificação de extratos vegetais obtidos de plantas brasileiras inibidores da enzima tirosinase, responsável pela biossíntese de melanina. Foi utilizada uma análise in vitro desenvolvida para se reproduzir a catalisação da L-tirosina a L-dopaquinona, um composto que apresenta coloração, detectável em espectrofotômetro em comprimento de onda de 490 nm. Foram testados 2.240 extratos vegetais orgânicos e aquosos, oriundos de plantas das florestas brasileiras. Os extratos foram testados em três concentrações diferentes, de 400, 200 e 100 ug/mL, e avaliados em quatro tempos, de 1h, 2h, 3h e 18h. Os extratos selecionados foram os que apresentaram atividade inibitória da tirosinase nas três concentrações e nos quatro tempos avaliados, o que resultou em 39 extratos ativos, correspondentes a 1,74% da quantidade total de extratos. Foram obtidos os valores referentes à concentração porcentagem de inibição enzimática e as curvas concentração x tempo. Os 39 extratos, relacionados segundo a eficácia em inibir a tirosinase, são os seguintes: N1229, N129, N2145, N471, N1809, N1497, N1415, N2014, N1959, N1879, N399, N1017, N135, N825, N257, N2247, N841, N1751, N861, N117, N1893, N59, N1873, N1323, N1224, N400, N137, N1365, N83, N503, N349, N1677, N1697, N1961, N215, N1414, N68, N1477, N1643. Ante o potencial terapêutico identificado nesses extratos, futuros estudos deverão ser feitos, de modo a se identificar os componentes ativos, examinar sua atividade quando incorporados em formulações dermocosméticas e verificar, por meio de testes in vitro, o grau de toxicidade que estes podem apresentar.

Pigmental disorders are dermal manifestations that change tonality of the skin and can result in an esthetic discomfort. Substances with a suppressing activity in the process of melanogenesis are used in the treatment of hyperchromia, especially inhibitors of the enzyme tyrosinase. The depigmentation substances that are currently used are not efficiently enough and safe for longer periods of time. New researches are necessary to find out which active inhibitors of melanogenesis are the strongest and safest ones. This research was made in order to identify plant extracts obtained from Brazilian plants that are inhibitors of the enzyme tyrosinase, responsible for melanin biosynthesis. The tests were made by in vitro analysis for the purpose of reproducing the L-tyrosine to L-dopaquinone, a compound that has a color detectable in a spectrophotometer at 490 nm. The amount of 2.240 organic and aqueous plant extracts were tested, all coming from Brazilian forests. The extracts were tested at three different concentrations of 400, 200 and 100ug/mL, and evaluated four times, during the period of 1h, 2h, 3h and 18h. The selected extracts were those that showed tyrosinase inhibitory activity in the three concentrations and during the four evaluated times, that resulted in 39 active extracts, corresponding to 1.74% of the amount of extracts. The values obtained were related to the percentage concentration of enzymatic inhibition and the concentration x time. All 39 extracts, that were organized according to the effectiveness in inhibiting tyrosinase, are: N1229, N129, N2145, N471, N1809, N1497, N1415, N2014, N1959, N1879, N399, N1017, N135, N825, N257, N2247, N841, N1751, N861, N117, N1893, N59, N1873, N1323, N1224, N400, N137, N1365, N83, N503, N349, N1677, N1697, N1961, N215, N1414, N68, N1477, N1643. As the therapeutic potential was identified in these extracts in the research future studies should be done, in order to identify the active components, verify their activity when incorporated in dermocosmetic formulations and verify, by means of in vitro tests, the degree of toxicity they may present.

Production and characterization of tyrosinase activity in Pycnoporus sanguineus CCT-4518 crude extract

Tyrosinase is an enzyme of industrial interest. The production and characterization of tyrosinase from P. sanguineus CCT-4518 were investigated. The selection of inductors, luminosity influence, inoculum size and type of culture medium on the production of tyrosinase and the effect of inhibitors on enzyme activity were performed. Optimum conditions for intracellular tyrosinase production was observed after 2 days using 0.15% L-tyrosine as inducer, in the presence of light, with inoculum size of 10 mycelium discs, using 2% malt extract broth medium, incubated at 30°C, and constant agitation of 150 rpm. Tyrosinase activity was completely inhibited by the addition of 6 mM salicylhydroxamic acid or phenylthiourea, however an inhibition of 4.15% was recorded by the addition of 0.1 mM sodium azide. No inhibition could be detected in case of 0.1 mM phenyl methanesulfonyl fluoride addition. Optimal conditions for intracellular tyrosinase activity using L-dopa as substrate were observed at pH 6.6 and 45°C. Thermal stability studies indicated that the enzyme is stable at 45°C for 15 minutes. Higher temperatures decreased tyrosinase activity. Enzyme production was confirmed by non-denaturing polyacrylamide gel electrophoresis and the protein profile was investigated by denaturing polyacrylamide gel electrophoresis.

Mutation of Aspergillus oryzae for improved production of 3, 4-dihydroxy phenyl-L-alanine (L-DOPA) from L-tyrosine

Aspergillus oryzae mutant strain UV-7 was further improved for the production of L-DOPA from L-tyrosine using chemical mutation. Different putative mutant strains of organism were tested for the production of L-DOPA in submerged fermentation. Among these putative mutant strains, mutant designated SI-12 gave maximum production of L-DOPA (300 mg L-DOPA.g-1 cells). The production of L-DOPA from different carbon source solutions (So= 30 g.l-1) by mutant culture was investigated at different nitrogen sources, initial pH and temperature values. At optimum pH (pHo= 5.0), and temperature (t=30ºC), 100% sugars were utilized for production and cell mass formation, corresponding to final L-DOPA product yield of 150 mg.g-1 substrate utilized, and maximum volumetric and specific productivities of 125 mg.l-1.h-1, and 150 mg.g-1 cells. h-1, respectively. There was up to 3-fold enhancement in product formation rate. This enhancement is the highest reported in literature. To explain the kinetic mechanism of L-DOPA formation and thermal inactivation of tyrosinase, the thermodynamic parameters were determined with the application of Arrhenius model: activation enthalpy and entropy for product formation, in case of mutant derivative, were 40 k j/mol and 0.076 k j/mol. K for L-DOPA production and 116 k j/mol and 0.590 k j/mol. K for thermal inactivation, respectively. The respective values for product formation were lower while those for product deactivation were higher than the respective values for the parental culture. Therefore, the mutant strain was thermodynamically more resistant to thermal denaturation.

A produção de L-DOPA a partir de tirosina pela cepa mutante de Aspergillus orizae UV-7 foi melhorada através de mutação química. Diferentes cepas foram testadas quanto a produção de L-DOPA por fermentação submersa, observando-se que a cepa denominada SI-12 foi a melhor produtora (300 mg de L-DOPA por g de células). A produção de L-DOPA pela cepa mutante a partir de diferentes fontes de carbono foi testada em diferentes fontes de nitrogênio, pH inicial e temperatura. Em pH ótimo (5,0) e temperatura ótima (30ºC), todos os açúcares foram utilizados para formação de biomassa, com um rendimento de L-DOPA de 150 mg.g-1, e produtividade volumétrica máxima e especifica de 125 mg.l.h-1 e 150 mg.g-1.h-1, respectivamente. A velocidade de formação do produto aumentou 3 vezes, sendo esse aumento o maior já relatado na literatura. Para explicar o mecanismo cinético da formação de L-DOPA e a inativação térmica da tirosinase, os parâmetros termodinâmicos foram determinados aplicando-se o modelo de Arrhenius: no caso da cepa mutante, a entalpia de ativação e entropia foram 40kj/mol e 0,076 kj/mol.K para produção de L-DOPA e 116 kj/mol and 0,590 kj/mol.K para inativação térmica, respectivamente. Os valores para formação do produto foram mais baixos e os para desativação do produto foram mais elevados que os valores correspondentes à cultura parental, indicando que a cepa mutante foi termodinamicamente mais resistente à denaturação térmica.

Melanoma maligno em búfalos (Bubalus bubalis) albinos / Malignant melanoma in albino water buffalo (Bubalus bubalis)

Descrevem-se melanomas múltiplos em dois búfalos albinos da raça Murrah no Sul do Brasil. Macroscopicamente havia múltiplos tumores enegrecidos no músculo esquelético, pulmões, linfonodos, saco pericárdio, cápsula renal, mediastino e pleura. Microscopicamente foram observadas células epitelióides poliédricas dispostas em ninhos sólidos ou fascículos entrelaçados sustentadas por um estroma colagenoso fino e esparso. O citoplasma era eosinofílico e por vezes continham quantidades variáveis de pigmento de melanina. A taxa de mitose foi baixa. Na imuno-histoquímica houve imunomarcação positiva utilizando anticorpo anti-tirosinase, anti-Melan-A, anti-vimentina, anti-proteína S-100 e antineurofilamento. É possível que o polimorfismo relacionado com o gene da pigmentação em búfalos albinos tenha contribuído para o maior risco de desenvolvimento de melanomas, como foi sugerido em humanos.

Two albino water buffalo affected by multiple melanocytic tumors in Southern Brazil are described. Grossly, there were multiple dark tumors within the skin, skeletal muscle, lungs, and lymph nodes. The tumor was also present in the pericardial sac, renal capsule, mediastinum and pleura. Microscopically, the tumors consisted of polyhedral epithelioid cells arranged in solid nests or interwoven fascicles supported by a thin and sparse collagenous stroma. The cytoplasm was eosinophilic and sometimes contained varying amounts of melanin pigment. The mitotic rates were low. Immunohistochemistry staining with Tyrosinase, Melan-A, Vimentin, S-100 protein, and neurofilament were positive. It is possible that the polymorphisms related to pigmentation in albino buffalo contributed with a high risk of developing melanoma as suggested in humans.

Melanoma maligno em búfalos (Bubalus bubalis) albinos / Malignant melanoma in albino water buffalo (Bubalus bubalis)

Descrevem-se melanomas múltiplos em dois búfalos albinos da raça Murrah no Sul do Brasil. Macroscopicamente havia múltiplos tumores enegrecidos no músculo esquelético, pulmões, linfonodos, saco pericárdio, cápsula renal, mediastino e pleura. Microscopicamente foram observadas células epitelióides poliédricas dispostas em ninhos sólidos ou fascículos entrelaçados sustentadas por um estroma colagenoso fino e esparso. O citoplasma era eosinofílico e por vezes continham quantidades variáveis de pigmento de melanina. A taxa de mitose foi baixa. Na imuno-histoquímica houve imunomarcação positiva utilizando anticorpo anti-tirosinase, anti-Melan-A, anti-vimentina, anti-proteína S-100 e antineurofilamento. É possível que o polimorfismo relacionado com o gene da pigmentação em búfalos albinos tenha contribuído para o maior risco de desenvolvimento de melanomas, como foi sugerido em humanos.

Two albino water buffalo affected by multiple melanocytic tumors in Southern Brazil are described. Grossly, there were multiple dark tumors within the skin, skeletal muscle, lungs, and lymph nodes. The tumor was also present in the pericardial sac, renal capsule, mediastinum and pleura. Microscopically, the tumors consisted of polyhedral epithelioid cells arranged in solid nests or interwoven fascicles supported by a thin and sparse collagenous stroma. The cytoplasm was eosinophilic and sometimes contained varying amounts of melanin pigment. The mitotic rates were low. Immunohistochemistry staining with Tyrosinase, Melan-A, Vimentin, S-100 protein, and neurofilament were positive. It is possible that the polymorphisms related to pigmentation in albino buffalo contributed with a high risk of developing melanoma as suggested in humans.

Melanoma maligno em búfalos (Bubalus bubalis) albinos / Malignant melanoma in albino water buffalo (Bubalus bubalis)

Descrevem-se melanomas múltiplos em dois búfalos albinos da raça Murrah no Sul do Brasil. Macroscopicamente havia múltiplos tumores enegrecidos no músculo esquelético, pulmões, linfonodos, saco pericárdio, cápsula renal, mediastino e pleura. Microscopicamente foram observadas células epitelióides poliédricas dispostas em ninhos sólidos ou fascículos entrelaçados sustentadas por um estroma colagenoso fino e esparso. O citoplasma era eosinofílico e por vezes continham quantidades variáveis de pigmento de melanina. A taxa de mitose foi baixa. Na imuno-histoquímica houve imunomarcação positiva utilizando anticorpo anti-tirosinase, anti-Melan-A, anti-vimentina, anti-proteína S-100 e antineurofilamento. É possível que o polimorfismo relacionado com o gene da pigmentação em búfalos albinos tenha contribuído para o maior risco de desenvolvimento de melanomas, como foi sugerido em humanos.

Two albino water buffalo affected by multiple melanocytic tumors in Southern Brazil are described. Grossly, there were multiple dark tumors within the skin, skeletal muscle, lungs, and lymph nodes. The tumor was also present in the pericardial sac, renal capsule, mediastinum and pleura. Microscopically, the tumors consisted of polyhedral epithelioid cells arranged in solid nests or interwoven fascicles supported by a thin and sparse collagenous stroma. The cytoplasm was eosinophilic and sometimes contained varying amounts of melanin pigment. The mitotic rates were low. Immunohistochemistry staining with Tyrosinase, Melan-A, Vimentin, S-100 protein, and neurofilament were positive. It is possible that the polymorphisms related to pigmentation in albino buffalo contributed with a high risk of developing melanoma as suggested in humans.