RESUMO
MafB is a transcription factor that regulates macrophage differentiation. Macrophages are a traditional feature of the hamster Harderian gland (HG); however, studies pertaining to MafB expression in the HG are scant. Here, the full-length cDNA of the MafB gene in hamsters was cloned and sequenced. Molecular characterization revealed that MafB encodes a protein containing 323 amino acids with a DNA-binding domain, a transactivation domain, and a leucine zipper domain. qPCR assays indicated that MafB was expressed in different tissues of both sexes. The highest relative expression levels in endocrine tissues were identified in the pancreas. Gonadectomy in male hamsters was associated with significantly higher mRNA levels in the HG; replacement with dihydrotestosterone restored mRNA expression. The HG in male hamsters contained twofold more MafB mRNA than the HG of female hamsters. Adrenals revealed similar mRNA relative expression levels during the estrous cycle. The estrous phase was associated with higher mRNA levels in the ovary. A significantly up-regulated expression and sexual dimorphism of MafB was found in the pancreas. Therefore, MafB in the HG may play an active role in the macrophage differentiation required for phagocytosis activity and intraocular repair. Additionally, sex steroids appear to strongly influence the MafB expression in the HG and pancreas. These studies highlight the probable biological importance of MafB in immunological defense and pancreatic ß cell regulation.
RESUMO
The molecular action of SOX9 can promote lipogenesis. Because the hamster Harderian gland (HG) synthesizes lipids and exhibits sexual dimorphism, this study aimed to identify and characterize Harderian SOX9. We examined the tissue distribution and expression profiles of SOX9 in hamster Mesocricetus auratus HGs. The full-length SOX9 cDNA sequence [3649-base pairs (bp)] contains an 81-bp 5' untranslated region (UTR), a 3' UTR of 2044-bp, an open reading frame (ORF) of 1524-bp, and a polyadenylation signal (AATAAA) at 19-bp upstream of poly(A) tail. The cDNA encodes a 507 amino acid protein containing the potential DNA-binding domain known as the HMG box. BLAST analysis revealed 99%, 99%, and 97% identity with the SOX9 of mouse, rat, and human, respectively. High expression levels were also observed in the testis, cerebellum, and hypothalamus. qPCR analysis demonstrated that SOX9 is expressed more abundantly in the HGs of males than in females. Sexually dimorphic expression of SOX9 suggests that differential expression between male and female HGs could be under the regulation of sex steroids. SOX9 might play a similar role in regulating exocrine secretions of lipids; these could occur downstream of FGF signaling - as found during embryogenesis - and/or androgen signaling.
Assuntos
Regulação da Expressão Gênica , Mesocricetus/metabolismo , Fatores de Transcrição SOX9/fisiologia , Animais , Biologia Computacional , Cricetinae , DNA Complementar/metabolismo , Feminino , Perfilação da Expressão Gênica , Lipogênese , Masculino , Conformação Molecular , Reação em Cadeia da Polimerase em Tempo Real , Fatores Sexuais , Distribuição TecidualRESUMO
The ATP-Binding Cassette, subfamily G, member 2 (ABCG2) transporter is associated with the regulation of protoporphyrin IX transport and of other intermediates in heme biosynthesis. Because the hamster Harderian gland (HG) exhibits high concentrations of porphyrins and sexual dimorphism, we analyzed the hamster ABCG2. Cloned cDNA [2098-base pairs (bp)] contains an open-reading frame (ORF) of 1971-bp that encodes a 656 amino-acid protein with a molecular weight of 72844.56Da. The hamster ABCG2 sequence is conserved phylogenetically and shares a high percentage of identity with mouse (89%), rat (88%), and human (84%) transporters. Within its structure, a Walker A (G-X-X-G-X-G-K-S), a C signature motif characteristic of ABC transporters, and six putative transmembrane domains (TMDs) were identified. ABCG2 mRNA was detected in all hamster tissues, with higher amounts found in HG, brain, cerebellum, kidney, gut, ovary, and testis. Harderian ABCG2 expression exhibits a sexually dimorphic pattern where females display higher mRNA levels than males. Different patterns of transcriptional profiles of ABCG2 during the estrous cycle and after gonadectomy in both sexes were also observed. The differential expression between male and female HGs suggests that ABCG2 is under the regulation of gonadal steroids. The ABCG2 transporter is likely involved in the endogenous regulation of porphyrins in hamster HGs.
Assuntos
Membro 2 da Subfamília G de Transportadores de Cassetes de Ligação de ATP/metabolismo , Hormônios Esteroides Gonadais/metabolismo , Glândula de Harder/metabolismo , Protoporfirinas/metabolismo , Animais , Cricetinae , DNA Complementar , Feminino , Humanos , Masculino , Mesocricetus , Camundongos , Porfirinas/metabolismo , RNA Mensageiro , Ratos , Caracteres SexuaisRESUMO
The objective of this study was to determine how cytokine transcription profiles correlate with patterns of infectious laryngotracheitis virus (ILTV) replication in the trachea, Harderian gland, and trigeminal ganglia during the early and late stages of infection after intratracheal inoculation. Viral genomes and transcripts were detected in the trachea and Harderian gland but not in trigeminal ganglia. The onset of viral replication in the trachea was detected at day one post-infection and peaked by day three post-infection. The peak of pro-inflammatory (CXCLi2, IL-1ß, IFN-γ) and anti-inflammatory (IL-13, IL-10) cytokine gene transcription, 5 days post-infection, coincided with the increased recruitment of inflammatory cells, extensive tissue damage, and limiting of virus replication in the trachea. In contrast, transcription of the IFN-ß gene in the trachea remained unaffected suggesting that ILTV infection blocks type I interferon responses. In the Harderian gland, the most evident transcription change was the early and transient upregulation of the IFN-γ gene at 1 day post-infection, which suggests that the Harderian gland is prepared to rapidly respond to ILTV infection. Overall, results from this study suggest that regulation of Th1 effector cells and macrophage activity by Th1/2 cytokines was pertinent to maintain a balanced immune response capable of providing an adequate Th1-mediated protective immunity, while sustaining some immune homeostasis in preparation for the regeneration of the tracheal mucosa.
Assuntos
Citocinas/metabolismo , Glândula de Harder/metabolismo , Infecções por Herpesviridae/veterinária , Herpesvirus Galináceo 1/patogenicidade , Traqueia/metabolismo , Gânglio Trigeminal/metabolismo , Animais , Galinhas , Citocinas/genética , DNA , Regulação da Expressão Gênica/imunologia , Genoma Viral , Glândula de Harder/virologia , Infecções por Herpesviridae/metabolismo , Infecções por Herpesviridae/virologia , Herpesvirus Galináceo 1/fisiologia , Doenças das Aves Domésticas/imunologia , Doenças das Aves Domésticas/metabolismo , Doenças das Aves Domésticas/virologia , RNA , Organismos Livres de Patógenos Específicos , Traqueia/virologia , Transcrição Gênica , Gânglio Trigeminal/virologia , Carga Viral , Virulência , Replicação ViralRESUMO
Protoporphyrin IX (PpIX) is a precursor of heme synthesis and is known to be an active photosensitizer and precursor of photosensitizers applied in photodynamic therapy (PDT) and photodynamic diagnostics (PDD). On irradiation with visible light, PpIX undergoes phototransformation, producing photoproducts which may also be phototoxic and increase its efficacy. The mechanism of PpIX phototransformation depends on environmental characteristics and can be different in vitro and in vivo. In this paper, we present a comparative study of the photoactivity of synthetic PpIX and PpIX extracted from the Harderian gland of ssp Rattus novergicus albinus rats, along with their photoproducts toward murine B16F-10 melanoma cells. It was observed that when irradiated with visible light the endogenous PpIX demonstrates photocytotoxicity ten times higher than the synthetic PpIX. The photoproduct of endogenous PpIX also possesses phototoxicity, though slightly lower than that of PpIX itself. The rate of cell internalization for both endogenous PpIX and its photoproduct was eightfold greater than that obtained for the synthetic porphyrin. This difference might result from a complexation of the native PpIX with some amphiphilic compounds during its synthesis within the Harderian glands, which facilitates the cell uptake of PpIX. Fluorescence microscopy images show that both endogenous and synthetic porphyrins are localized after uptake predominantly in the mitochondrial region of cells.
Assuntos
Melanoma/patologia , Fotoquimioterapia , Fármacos Fotossensibilizantes/farmacologia , Protoporfirinas/farmacologia , Animais , Transporte Biológico , Linhagem Celular Tumoral , Sobrevivência Celular/efeitos dos fármacos , Sobrevivência Celular/efeitos da radiação , Escuridão , Glândula de Harder/metabolismo , Espaço Intracelular/metabolismo , Masculino , Melanoma/tratamento farmacológico , Camundongos , Fármacos Fotossensibilizantes/síntese química , Fármacos Fotossensibilizantes/isolamento & purificação , Fármacos Fotossensibilizantes/metabolismo , Protoporfirinas/síntese química , Protoporfirinas/isolamento & purificação , Protoporfirinas/metabolismo , RatosRESUMO
The aim of the present study was to demonstrate the histological and histochemical structure of the Harderian gland in wild and hybrid of wild and domestic birds. The samples were stained with haematoxylin-eosin, methyl green-pyronin Y, periodic acid-Schiff, alcian blue pH 2.5, aldehyde fuchsin and Hale's dialyzed iron staining's. In both species, the glands had multilobar tubuloacinar structure type I. The Harderian gland was located in the orbit near the inter-orbital septum, between the medial rectus muscle, the pyramidal muscle of the third eyelid, and the ventral oblique muscle. In the common pheasant, the gland was wider in the proximal and distal part. The common pheasant had more elongated lobes of the Harderian gland than in the hybrid. In the common pheasant, the glandular cells presented darkly-stained serous secretion and lightly-stained mucous secretion. In the hybrid, the glandular cells presented seromucous secretion. The central lobar space, interacinar space, and apical parts of the acini of the Harderian glands were filled with many lymphocytes and plasma cells, particularly in the common pheasant, where centers of all large lobes were abundantly filled with plasma cells. The plasma cells dominated in common pheasant's Harderian gland, while in the hybrid, lymphocytes and plasma cells were present at similar quantities. The cells positive for periodic acid of Schiff staining were dominant in hybrid. Periodic acid-Schiff, Hale's dialyzed iron and alcian blue pH 2.5 stainings demonstrated acid-carboxylated mucopolysaccharides in the glandular cells cytoplasm of the examined birds.
Assuntos
Animais , Aves/anatomia & histologia , Aves/classificação , Glândula de Harder/anatomia & histologiaRESUMO
The aim of the present study was to demonstrate the histological and histochemical structure of the Harderian gland in wild and hybrid of wild and domestic birds. The samples were stained with haematoxylin-eosin, methyl green-pyronin Y, periodic acid-Schiff, alcian blue pH 2.5, aldehyde fuchsin and Hale's dialyzed iron staining's. In both species, the glands had multilobar tubuloacinar structure type I. The Harderian gland was located in the orbit near the inter-orbital septum, between the medial rectus muscle, the pyramidal muscle of the third eyelid, and the ventral oblique muscle. In the common pheasant, the gland was wider in the proximal and distal part. The common pheasant had more elongated lobes of the Harderian gland than in the hybrid. In the common pheasant, the glandular cells presented darkly-stained serous secretion and lightly-stained mucous secretion. In the hybrid, the glandular cells presented seromucous secretion. The central lobar space, interacinar space, and apical parts of the acini of the Harderian glands were filled with many lymphocytes and plasma cells, particularly in the common pheasant, where centers of all large lobes were abundantly filled with plasma cells. The plasma cells dominated in common pheasant's Harderian gland, while in the hybrid, lymphocytes and plasma cells were present at similar quantities. The cells positive for periodic acid of Schiff staining were dominant in hybrid. Periodic acid-Schiff, Hale's dialyzed iron and alcian blue pH 2.5 stainings demonstrated acid-carboxylated mucopolysaccharides in the glandular cells cytoplasm of the examined birds.(AU)
Assuntos
Animais , Aves/anatomia & histologia , Aves/classificação , Glândula de Harder/anatomia & histologiaRESUMO
According to current knowledge, two steroid 5α-reductases, designated type 1 (SRD5A1) and type 2 (SRD5A2), are present in all species examined to date. These isozymes play a central role in steroid hormone physiology by catalyzing the reduction of 3-keto-4-ene-steroids into more active 5α-reduced derivatives, including the conversion of testosterone (T) to dihydrotestosterone (DHT). A third 5α-reductase (SRD5A3, -type 3), which is overexpressed in hormone-refractory prostate cancer cells, has been identified; however, its enzymatic characteristics are practically unknown. Here, we isolated a cDNA encoding hamster Srd5a3 (hSrd5a3) and performed functional metabolic assays to investigate its biochemical properties. The cloned cDNA encodes a 330 amino acid protein that is 87% identical to the homologous protein in mice and 78% to that in humans. However, hSrd5a3 exhibits low sequence homology with its counterparts hSrd5a1 (19%) and hSrd5a2 (17%). A fusion protein consisting of hSrd5a3 and green fluorescent protein provided evidence for cytoplasmic localization in transfected mammalian cells. Real-time PCR analysis revealed that, Srd5a3 mRNA was present in nearly all hamster tissues, with high expression in the cerebellum, Harderian gland and testis. Functional assays expressing hSrd5a3 cDNA in HEK-293 cells revealed that this isozyme is unable to reduce T into DHT. Further expression assays confirmed that similar to testosterone, progesterone, androstenedione and corticosterone are not reduced by hSrd5a3 or human SRD5A3. Together, these results indicate that hSrd5a3 lacks the catalytic activity to transform 3-keto-4-ene-compounds; therefore 5α-reductase type 3 may not be involved in 5α-reduction of steroids.
Assuntos
3-Oxo-5-alfa-Esteroide 4-Desidrogenase/fisiologia , 3-Oxo-5-alfa-Esteroide 4-Desidrogenase/química , Sequência de Aminoácidos , Animais , Sequência de Bases , Biocatálise , Sequência Conservada , Cricetinae , Di-Hidrotestosterona/química , Feminino , Células HEK293 , Humanos , Masculino , Mesocricetus , Dados de Sequência Molecular , Especificidade de Órgãos , OxirreduçãoRESUMO
Purpose: To show that a non-laser light source can be constructed, using a 500 W Tungsten lamp and optical filters, and that this light source produces photodynamic effect via protoporphyrin IX (PpIX) similar to the effect produced by HeNe laser light. Methods: The broad band spectrum from a Tungsten lamp was filtered. Infrared and blue part of the spectrum was discarded by absorption process and the fraction of the spectrum, centered at the red portion, was filtered by an interference filter. Photodynamic effect was studied by the activity on endogenous PpIX of Harderian glands of Wistar rats. Twenty rats were used for the experiment. Each animal had its two Harderian gland surgically exposed, so one of them was treated with the system while the other was kept as control. After a 30 minutes period of treatment, the animals were sacrificed and their glands were removed for histological analysis. This analysis was compared to earlier published results obtained with HeNe laser light. Results: The resultant light source emission was centered around (636 ± 6.5) nm and gives up to 11.3 mW/cm² power density. It produces photodynamic effect in Harderian gland, observed either by fluorescence spectroscopy or by histological microscopy. Conclusion: There is no noticeable difference in Photodynamic effect results if activated by HeNe laser or by the proposed non-laser light source emitting at the red portion of the spectrum.
Objetivo: Descrever a construção de uma fonte de luz não-laser, a partir de uma lâmpada de Tungstênio e filtros óticos adequados e demonstrar que sua eficiência em estudos fotodinâmicos, mediados por protoporfirina IX, é semelhante a do laser de Hélio Neônio. Métodos: As regiões Infravermelha e Azul do espectro de emissão óptica de uma lâmpada de Tungstênio foram convenientemente descartadas por processos de absorção, enquanto que a fração centrada na região do vermelho foi removida com o uso de um filtro de interferência. O efeito fotodinâmico foi estudado, em glândula Harderiana de ratos Wistar em razão da produção endógena de protoporfirina IX (PpIX) por estas glândulas. Foram utilizados 20 ratos. Cada animal teve as duas glândulas expostas cirurgicamente, sendo uma delas tratada com a fonte não-laser e a outra mantida como controle. Após tratamento por 30 minutos os animais foram sacrificados e suas glândulas removidas para estudo histológico. Os resultados foram comparados a estudos realizados com laser de Hélio Neônio, já publicados. Resultados: A luz produzida pelo equipamento está centrada em torno de (636 ± 6,5) nm, fornecendo uma densidade de potência de 11,3 mW/cm². Os efeitos fotodinâmicos produzidos na glândula Harderiana, podem ser observados tanto por espectroscopia de fluorescência como por microscopia ótica. Conclusão: Não foram observadas diferenças significativas nos resultados do efeito fotodinâmico obtidos com a fonte de luz não-laser proposta, em comparação aos resultados conhecidos com o uso do laser de Hélio Neônio.