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1.
Sheng Wu Gong Cheng Xue Bao ; 36(8): 1590-1599, 2020 Aug 25.
Artigo em Chinês | MEDLINE | ID: mdl-32924357

RESUMO

ß-defensin is a primary protein immune factor in channel catfish's (Ietalurus punetaus) resistance to pathogenic microorganisms. Its primary structure contains a signal peptide composed of 24 amino acid residues at the N-terminal and a mature peptide composed of 43 amino acid residues at the C-terminal. The mature peptide region is responsible for the biological activity of ß-defensin. In the present study, a recombinant strain of Pichia pastoris that produces channel catfish ß-defensin, was constructed to realize the biosynthesis of channel catfish ß-defensin based on eukaryotic expression. First, the ß-defensin gene "IPBD" was isolated from the skin of channel catfish by RT-PCR. After linking it with the expression vector pPICZA, pPICZA-IPBD was transferred into competent P. pastoris X-33 cells to obtain recombinant P. pastoris strains. The yeast transformants with multi-copy gene inserts were obtained by using the culture medium containing 1 000 µg/mL zeocin. Using BMM culture medium (without amino nitrogen culture medium) instead of BMMY culture medium (with amino nitrogen culture medium), the fermentation and culture conditions of the recombinant strain were optimized, and the optimal conditions for producing channel catfish ß-defensin were determined as follows: the expression was induced for 96 h with 1.0% methanol at 28 °C , 250 r/min. Purified protein with molecular weight of 5.98 kDa was obtained by nickel affinity chromatography, and MALDI-TOF/TOF mass spectrometry proved that it was the expected recombinant IPBD. The antibacterial test results showed that the inhibitory rates of recombinant IPBD on Gram-positive Staphylococcus aureus and Listeria monocytogenes and Gram-negative Pseudomonas aeruginosa were 69.6%, 71.6% and 65.8%, respectively. This study provides a recombinant DNA technique for the development of small molecule natural antibacterial peptide from fish.


Assuntos
Expressão Gênica , Pichia , beta-Defensinas , Animais , Antibacterianos/farmacologia , Bactérias/efeitos dos fármacos , Ictaluridae/genética , Pichia/genética , beta-Defensinas/genética , beta-Defensinas/isolamento & purificação , beta-Defensinas/metabolismo , beta-Defensinas/farmacologia
2.
Arch Oral Biol ; 117: 104827, 2020 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-32673820

RESUMO

OBJECTIVE: The aim of the study was to assess whether or not Vitamin D deficiency was associated with the GCF and gingival tissue antimicrobial peptides (AMP), namely, human beta defensin-2 (hBD-2) and cathelicidin (LL-37) level in chronic periodontitis (CP) and gingivitis patients. DESIGN: A total of 80 volunteers were included in this study. Forty was classified as Vitamin D deficient (25(OH)D <20 ng/mL), and 40 Vitamin D sufficient patients (25(OH)D ≥ 20 ng/mL). Of these, 20 of them were affected by gingivitis and 20 by CP. Following sampling, the hBD-2 and LL-37 concentration in gingival tissues and GCF were determined by the ELISA method. RESULTS: The hBD-2 and LL-37 levels were higher in periodontitis compared to gingivitis patients within Vitamin D sufficient and deficient groups. The AMP levels of GCF and gingival tissue in the vitamin D deficient group was lower compared to sufficient serum 25(OH)D within gingivitis and CP groups. Additionally, a non-parametric regression model known as the generalized additive model was used to identify the contribution of diagnosis, Vitamin D status, and other potential clinical variables on the local levels of AMPs. Regression analysis showed that the periodontal disease status, serum vitamin D concentration were independent predictors for elevated GCF AMP levels. Finally, a positive correlation between GCF and tissue levels of both hBD-2 (r = 0.82; <0.0001) and LL-37 (r = 0.65; <0.0001) was detected. CONCLUSION: This study shows that serum 25(OH)D deficiency is associated with decreased hBD-2 and LL-37 expression of GCF and gingival tissues in both gingivitis and CP patients.


Assuntos
Peptídeos Catiônicos Antimicrobianos/metabolismo , Periodontite Crônica , Gengivite , Vitamina D/sangue , beta-Defensinas/metabolismo , Líquido do Sulco Gengival , Humanos
3.
Nat Commun ; 11(1): 2703, 2020 06 01.
Artigo em Inglês | MEDLINE | ID: mdl-32483147

RESUMO

Mother's milk is the best choice for infants nutrition, however when it is not available or insufficient to satisfy the needs of the infant, formula is proposed as an effective substitute. Here, we report the results of a randomized controlled clinical trial (NCT03637894) designed to evaluate the effects of two different dietary regimens (standard formula and Lactobacillus paracasei CBA L74-fermented formula) versus breastfeeding (reference group) on immune defense mechanisms (primary endpoint: secretory IgA, antimicrobial peptides), the microbiota and its metabolome (secondary outcomes), in healthy full term infants according to the type of delivery (n = 13/group). We show that the fermented formula, safe and well tolerated, induces an increase in secretory IgA (but not in antimicrobial peptides) and reduces the diversity of the microbiota, similarly, but not as much as, breastmilk. Metabolome analysis allowed us to distinguish subjects based on their dietary regimen and mode of delivery. Together, these results suggest that a fermented formula favors the maturation of the immune system, microbiota and metabolome.


Assuntos
Dieta , Sistema Imunitário/fisiologia , Metaboloma/fisiologia , Microbiota/fisiologia , Peptídeos Catiônicos Antimicrobianos/metabolismo , Aleitamento Materno , Método Duplo-Cego , Fezes/microbiologia , Feminino , Fermentação , Humanos , Imunoglobulina A Secretora/metabolismo , Fórmulas Infantis , Recém-Nascido , Lactobacillus paracasei/metabolismo , Masculino , Leite Humano , beta-Defensinas/metabolismo
4.
Eur J Histochem ; 64(2)2020 Apr 21.
Artigo em Inglês | MEDLINE | ID: mdl-32363846

RESUMO

Sperm-associated antigen 11A (SPAG11A) is expressed exclusively in the epididymis, which can specifically regulate sperm maturation. The aim of this study was to investigate the seasonal expressions of beta-defensin (SPAG11A) and androgen receptor (AR) in the epididymis of the wild ground squirrels (Citellus dauricus Brandt). Morphologically, the results showed that epididymis length and weight in the breeding season were significantly higher than those of the non-breeding season. Histologically, the results revealed that enlarged lumen diameters, thickened epithelial and abundant sperm in the breeding season while reduced lumen diameters and epithelial with no sperm in the non-breeding season. SPAG11A was intensely expressed in cytoplasm and nucleus of epithelial cells in the breeding season, and weaker staining in the non-breeding season. The immunostaining of AR was only presented in nucleus of smooth muscle cells and epithelial cells in the whole epididymis with stronger staining in the breeding season. The results of real-time quantitative PCR also showed that the mRNA levels of SPAG11A and AR in the epididymis during the breeding season were significantly higher than those of the non-breeding season. In addition, the levels of testosterone (T), follicle-stimulating hormone (FSH) and luteinizing hormone (LH) in the serum were higher during the breeding season. Taken together, these results suggested that SPAG11A might play an important role to regulate seasonal changes in epididymal function of the wild ground squirrels.


Assuntos
Epididimo/metabolismo , Receptores Androgênicos/metabolismo , Estações do Ano , beta-Defensinas/metabolismo , Animais , Cruzamento , Epididimo/citologia , Hormônio Foliculoestimulante/metabolismo , Imuno-Histoquímica , Hormônio Luteinizante/metabolismo , Masculino , Sciuridae , Testosterona/metabolismo
5.
Int J Oral Sci ; 12(1): 13, 2020 04 29.
Artigo em Inglês | MEDLINE | ID: mdl-32350241

RESUMO

Efforts to control inflammation and achieve better tissue repair in the treatment of periodontitis have been ongoing for years. Human ß-defensin 3, a broad-spectrum antimicrobial peptide has been proven to have a variety of biological functions in periodontitis; however, relatively few reports have addressed the effects of human periodontal ligament cells (hPDLCs) on osteogenic differentiation. In this study, we evaluated the osteogenic effects of hPDLCs with an adenoviral vector encoding human ß-defensin 3 in an inflammatory microenvironment. Then human ß-defensin 3 gene-modified rat periodontal ligament cells were transplanted into rats with experimental periodontitis to observe their effects on periodontal bone repair. We found that the human ß-defensin 3 gene-modified hPDLCs presented with high levels of osteogenesis-related gene expression and calcium deposition. Furthermore, the p38 MAPK pathway was activated in this process. In vivo, human ß-defensin 3 gene-transfected rat PDLCs promoted bone repair in SD rats with periodontitis, and the p38 mitogen-activated protein kinase (MAPK) pathway might also have been involved. These findings demonstrate that human ß-defensin 3 accelerates osteogenesis and that human ß-defensin 3 gene modification may offer a potential approach to promote bone repair in patients with periodontitis.


Assuntos
Anti-Infecciosos/farmacologia , Osteogênese/efeitos dos fármacos , Ligamento Periodontal/metabolismo , Periodontite/tratamento farmacológico , beta-Defensinas/farmacologia , Animais , Anti-Infecciosos/metabolismo , Diferenciação Celular/efeitos dos fármacos , Células Cultivadas , Humanos , Ligamento Periodontal/efeitos dos fármacos , Ratos , Ratos Sprague-Dawley , beta-Defensinas/metabolismo
6.
PLoS One ; 15(4): e0230231, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32240190

RESUMO

Enteroids are cultured primary intestinal epithelial cells that recapitulate epithelial lineage development allowing for a more complex and physiologically relevant model for scientific study. The large presence of intestinal stem cells (ISC) in these enteroids allows for the study of metabolite effects on cellular processes and resulting progeny cells. Short-chain fatty acids (SCFA) such as butyrate (BUT) are bacterial metabolites produced in the gastrointestinal tract that are considered to be beneficial to host cells. Therefore, the objective was to study the effects of SCFAs on biomarkers of ISC activity, differentiation, barrier function and epithelial defense in the intestine using mouse and human enteroid models. Enteroids were treated with two concentrations of acetate (ACET), propionate (PROP), or BUT for 24 h. Enteroids treated with BUT or PROP showed a decrease in proliferation via EdU uptake relative to the controls in both mouse and human models. Gene expression of Lgr5 was shown to decrease with BUT and PROP treatments, but increased with ACET. As a result of BUT and PROP treatments, there was an increase in differentiation markers for enterocyte, Paneth, goblet, and enteroendocrine cells. Gene expression of antimicrobial proteins Reg3ß, Reg3γ, and Defb1 were stimulated by BUT and PROP, but not by ACET which had a greater effect on expression of tight junction genes Cldn3 and Ocln in 3D enteroids. Similar results were obtained with human enteroids treated with 10 mM SCFAs and grown in either 3D or Transwell™ model cultures, although tight junctions were influenced by BUT and PROP, but not ACET in monolayer format. Furthermore, BUT and PROP treatments increased transepithelial electrical resistance after 24 h compared to ACET or control. Overall, individual SCFAs are potent stimulators of cellular gene expression, however, PROP and especially BUT show great efficacy for driving cell differentiation and gene expression.


Assuntos
Ácido Acético/farmacologia , Ácido Butírico/farmacologia , Regulação da Expressão Gênica/efeitos dos fármacos , Propionatos/farmacologia , Esferoides Celulares/efeitos dos fármacos , Animais , Técnicas de Cultura de Células , Diferenciação Celular/efeitos dos fármacos , Claudina-3/genética , Claudina-3/metabolismo , Enterócitos/citologia , Enterócitos/efeitos dos fármacos , Enterócitos/metabolismo , Células Enteroendócrinas/citologia , Células Enteroendócrinas/efeitos dos fármacos , Células Enteroendócrinas/metabolismo , Células Caliciformes/citologia , Células Caliciformes/efeitos dos fármacos , Células Caliciformes/metabolismo , Humanos , Camundongos , Ocludina/genética , Ocludina/metabolismo , Proteínas Associadas a Pancreatite/genética , Proteínas Associadas a Pancreatite/metabolismo , Celulas de Paneth/citologia , Celulas de Paneth/efeitos dos fármacos , Celulas de Paneth/metabolismo , Cultura Primária de Células , Receptores Acoplados a Proteínas-G/genética , Receptores Acoplados a Proteínas-G/metabolismo , Esferoides Celulares/citologia , Esferoides Celulares/metabolismo , Junções Íntimas/efeitos dos fármacos , beta-Defensinas/genética , beta-Defensinas/metabolismo
7.
Anim Sci J ; 91(1): e13372, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32285528

RESUMO

Antibacterial factors act as innate immune components, which respond as soon as bacteria enter a living organism. To prevent and treat mastitis in cattle, understanding the concentrations of these substances inside the udder is important; however, they remain to be studied. In this investigation, the concentration of lingual antimicrobial peptide (LAP), S100 protein (S100A7), lactoferrin (LF), and immunoglobulin antibody were measured in the different fractions of foremilk. Lactating Holstein cows were examined, and 10 foremilk fractions were obtained from sequential samples up to 150 ml. The LAP concentrations in milk samples increased until 25 ml. The LF concentrations increased up to the 10 ml fraction, then stabilized at low level after the 50 ml fraction. For S100A7, some fractions had significantly higher (p < .05) concentrations than the 5 or 10 ml fractions. The IgA antibody concentration increased up to the 5 ml fraction, then after 50 ml fraction showed relatively low concentrations. This investigation determined the concentration patterns of LAP, LF, S100A7, and IgA antibody secreted in milk inside the udders of healthy lactating cows as baseline data. These distinct concentration patterns might indicate various protective responses.


Assuntos
Imunoglobulina A/metabolismo , Lactoferrina/metabolismo , Leite/imunologia , Leite/metabolismo , Proteína A7 Ligante de Cálcio S100/metabolismo , beta-Defensinas/metabolismo , Animais , Bovinos , Feminino , Imunidade Inata , Lactação , Glândulas Mamárias Animais/metabolismo
8.
Anim Sci J ; 91(1): e13365, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32285581

RESUMO

The present study was conducted to examine whether colostrum supplementation in peripartum goats increases the antimicrobial peptides in their milk. Goats were orally administered 2 ml of colostrum whey products (colostrum group) or water (control group) daily, from 2 weeks before until 2 weeks after kidding. Body weights of mothers and kids were measured. Blood, milk, and fecal samples were collected from the mothers, and blood samples were collected from the kids. Concentrations of milk antimicrobial peptides (beta-defensin, cathelicidin, lactoferrin, S100A7, lactoperoxidase, and immunoglobulin A [IgA]) were determined. IgA and nutritional parameters (glucose, total cholesterol, triglyceride, ketone bodies, and non-esterified fatty acids) were also determined in the blood of mothers and kids. Milk IgA and lactoferrin concentrations were higher in the colostrum group than in the control group. Conversely, lower milk concentrations of S100A7 were observed in the colostrum group than that in the control group. Plasma IgA concentrations were higher for kids from the colostrum group than for those from the control group. These results suggest that oral administration of colostrum in pregnant goats increases IgA concentration in postpartum milk, which can subsequently improve the health of their kids.


Assuntos
Fenômenos Fisiológicos da Nutrição Animal , Peptídeos Catiônicos Antimicrobianos/metabolismo , Colostro , Dieta/veterinária , Suplementos Nutricionais , Lactoferrina/metabolismo , Leite/metabolismo , Proteínas do Soro do Leite/administração & dosagem , beta-Defensinas/metabolismo , Administração Oral , Animais , Feminino , Cabras , Imunoglobulina A/metabolismo , Lactoperoxidase/metabolismo , Período Periparto , Gravidez
9.
PLoS One ; 15(3): e0229647, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32150574

RESUMO

Probiotic bacteria have the ability to modulate host immune responses and have potent therapeutic functional effects against several diseases, including inflammatory diseases. However, beneficial effects of probiotics are strain specific and their interactions with host immune cells to modulate inflammatory response are largely unknown. Intestinal epithelial cells (IECs), which are the first line of defense against invading pathogens, and connects between commensals/probiotics and immune system; therefore, in this study, we used human IECs to assess the probiotic effects of three selected Lactobacillus strains in vitro. An HT-29 colonic epithelial cell and HT-29/blood mononuclear cells co-culture system were stimulated with Lactobacillus followed by Salmonella for different hours, after which the mRNA level of cytokines, ß-defensin-2 and negative regulators for TLR signaling and protein levels of ZO-1 and IκB-α were analyzed by real-time polymerase chain reaction and western blot analysis. L. brevis decreased Salmonella induced IL-6, IL-8, MCP-1 and IL-1ß levels, whereas L. pentosus suppressed IL-6 and MCP-1 in HT-29 cells. Moreover, L. brevis was able to increase the mRNA levels of A20, Tollip, SIGIRR and IRAKM, while L. pentosus reduced the levels of A20, and IRAKM in response to Salmonella. In addition, decrease in protein level of TNF-α and increase in mRNA level of IL-10 was observed in L. brevis and L. pentosus treated HT-29 cells. Lactobacillus strains were differentially modulated ZO-1 and p-IκB-α in HT-29 cells treated with Salmonella. Overall, the results of this study indicate that Lactobacillus strains attenuate Salmonella induced inflammatory responses through beneficial modulation of TLR negative regulators and the NF-κB pathway.


Assuntos
Mucosa Intestinal/imunologia , Mucosa Intestinal/microbiologia , Probióticos/uso terapêutico , Salmonella/imunologia , Salmonella/patogenicidade , Técnicas de Cocultura , Citocinas/metabolismo , Células Epiteliais/imunologia , Células Epiteliais/microbiologia , Células HT29 , Interações entre Hospedeiro e Microrganismos/imunologia , Humanos , Lactobacillus/imunologia , Leucócitos Mononucleares/imunologia , Leucócitos Mononucleares/microbiologia , NF-kappa B/metabolismo , Infecções por Salmonella/imunologia , Infecções por Salmonella/prevenção & controle , Transdução de Sinais , Receptores Toll-Like/metabolismo , beta-Defensinas/metabolismo
10.
Anal Biochem ; 594: 113627, 2020 04 01.
Artigo em Inglês | MEDLINE | ID: mdl-32067984

RESUMO

We used base-appended base modification to develop a new adenine analog, which incorporates an adenine derivative at position 7 of adenine. Using the systematic evolution of ligands by exponential enrichment method with a modified DNA library including this analog, we obtained Aad1, an aptamer that binds strongly to human ß-defensin 2, a biomarker of physical stress found in saliva. The dissociation constant of Aad1 with respect to human ß-defensin 2 was found to be low (6.8 nM), and was found to bind specifically to human ß-defensin 2 in saliva spiked with the protein, as confirmed using pull-down with magnetic beads. To our knowledge, there are no prior reports of nucleic-acid aptamers that bind specifically to human ß-defensin 2. However, our results indicated that such adenine analog-containing DNA libraries are extremely effective in the acquisition of high-affinity aptamers.


Assuntos
Adenina/análogos & derivados , Aptâmeros de Nucleotídeos/metabolismo , Técnica de Seleção de Aptâmeros/métodos , beta-Defensinas/metabolismo , Humanos , Ligação Proteica , Saliva/metabolismo
11.
PLoS One ; 15(2): e0228656, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32053710

RESUMO

Fresh semen is most commonly used in an artificial insemination of small ruminants, because of low fertility rates of frozen sperm. Generally, when developing and applying assisted reproductive technologies, sheep and goats are classified as one species. In order to optimize sperm cryopreservation protocols in sheep and goat, differences in sperm proteomes between ram and buck are necessary to investigate, which may contribute to differences in function and fertility of spermatozoa. In the current work, a data-independent acquisition-mass spectrometry proteomic approach was used to characterize and make a comparison of ram (Ovis aries) and buck (Capra hircus) sperm proteomes. A total of 2,109 proteins were identified in ram and buck spermatozoa, with 238 differentially abundant proteins. Proteins identified in ram and buck spermatozoa are mainly involved in metabolic pathways for generation of energy and diminishing oxidative stress. Specifically, there are greater abundance of spermatozoa proteins related to the immune protective and capacity activities in ram, while protein that inhibit sperm capacitation shows greater abundance in buck. Our results not only provide novel insights into the characteristics and potential activities of spermatozoa proteins, but also expand the potential direction for sperm cryopreservation in ram and buck.


Assuntos
Proteoma , Espermatozoides/metabolismo , Animais , Proteína C-Reativa/metabolismo , Criopreservação , Complexo IV da Cadeia de Transporte de Elétrons/metabolismo , Cabras , Concentração de Íons de Hidrogênio , Masculino , Espectrometria de Massas , Proteínas do Tecido Nervoso/metabolismo , Estresse Oxidativo , Análise de Componente Principal , Mapeamento de Interação de Proteínas , Proteômica , Preservação do Sêmen , Ovinos , Capacitação Espermática , Motilidade Espermática , Succinato-Semialdeído Desidrogenase/metabolismo , beta-Defensinas/metabolismo
12.
Genet Test Mol Biomarkers ; 24(3): 113-119, 2020 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-32058800

RESUMO

Background: Human ß-defensin-2 is an antimicrobial peptide with antibiotic properties secreted by the oral cavity to protect the host against microbial attack. The inter-individual differences in defensin expression profiles due to genetic variation might be partly responsible for differences in disease susceptibility. Aims: The objective of this study was to examine whether variation in the human ß-defensin-2 gene (DEFB4A) is associated with chronic periodontitis (CP). Materials and Methods: This case-control study used Sanger sequencing to analyze two promoter polymorphisms of the DEFB4A gene with potential functional consequences using DNA samples collected from 200 unrelated individuals. Results: The DEFB4A rs1339258595 promoter polymorphism is associated with CP risk and clinical attachment level (CAL) but the rs3762040 polymorphism is not. Carriers of the T allele (rs1339258595) were approximately three times less likely to develop periodontitis compared with noncarriers (p = 0.0004, odds ratio = 0.35). Consistent with a protective role, the carriers of T allele had a lower CAL compared with the wild-type (G) allele. Moreover, the wild-type diplotype (GGGG) had a significantly higher risk of tooth loss compared with other diplotypes (p = 0.016). Conclusion: This study demonstrates that genetic variation in the promoter region of DEFB4A likely affects resistance to periodontal infection and might be a potential marker for CP risk and severity.


Assuntos
Periodontite Crônica/genética , beta-Defensinas/genética , Adulto , Alelos , Estudos de Casos e Controles , Suscetibilidade a Doenças , Feminino , Frequência do Gene/genética , Predisposição Genética para Doença/genética , Genótipo , Heterozigoto , Humanos , Masculino , Pessoa de Meia-Idade , Razão de Chances , Polimorfismo de Nucleotídeo Único/genética , Regiões Promotoras Genéticas/genética , Turquia , beta-Defensinas/metabolismo
13.
Artigo em Inglês | MEDLINE | ID: mdl-32092464

RESUMO

Patients with Atopic Dermatitis (AD) suffer from inflamed skin and skin barrier defects. Proper formation of the outermost part of the skin, the stratum corneum (SC), is crucial for the skin barrier function. In this study we analyzed the localization and activity of lipid enzymes ß-glucocerebrosidase (GBA) and acid sphingomyelinase (ASM) in the skin of AD patients and controls. Localization of both the expression and activity of GBA and ASM in the epidermis of AD patients was altered, particularly at lesional skin sites. These changes aligned with the altered SC lipid composition. More specifically, abnormal localization of GBA and ASM related to an increase in specific ceramide subclasses [AS] and [NS]. Moreover we related the localization of the enzymes to the amounts of SC ceramide subclasses and free fatty acids (FFAs). We report a correlation between altered localization of active GBA and ASM and a disturbed SC lipid composition. Localization of antimicrobial peptide beta-defensin-3 (HBD-3) and AD biomarker Thymus and Activation Regulated Chemokine (TARC) also appeared to be diverging in AD skin compared to control. This research highlights the relation between correct localization of expressed and active lipid enzymes and a normal SC lipid composition for a proper skin barrier.


Assuntos
Dermatite Atópica/imunologia , Epiderme/patologia , Glucosilceramidase/metabolismo , Metabolismo dos Lipídeos/imunologia , Esfingomielina Fosfodiesterase/metabolismo , Adolescente , Adulto , Biópsia , Estudos de Casos e Controles , Ceramidas/análise , Ceramidas/metabolismo , Quimiocina CCL17/metabolismo , Dermatite Atópica/patologia , Epiderme/química , Epiderme/enzimologia , Ácidos Graxos não Esterificados/análise , Ácidos Graxos não Esterificados/metabolismo , Feminino , Voluntários Saudáveis , Humanos , Masculino , Perda Insensível de Água/imunologia , Adulto Jovem , beta-Defensinas/metabolismo
14.
Res Vet Sci ; 129: 28-38, 2020 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-31927467

RESUMO

The ovine rumen is an immune interface with the external environment, participating in host defence responses. Ovine ruminal epithelial cells (ORECs) not only have a physical barrier function, but also secrete sheep ß-defensin-1 (SBD-1), which plays a key role in innate and adaptive immunity. Prebiotics are potential alternatives to infeed antibiotics. Saccharomyces cerevisiae cell wall (S.c.CW) is rich in prebiotics, which play roles in improving the growth performance of animals and regulating immunity. Here, we investigated whether S.c.CW induces SBD-1 expression in ORECs, as well as the underlying mechanism. The regulatory mechanisms of S.c.CW-induced up-regulation of SBD-1 were determined using quantitative real-time PCR, enzyme-linked immunosorbent assay, and western blotting. S.c.CW significantly increased the expression of Toll-like receptor 2 (TLR2) and nuclear factor-kappa B (NF-κB), but had no effect on TLR4 expression. TLR2, MyD88, and NF-κB inhibition attenuated the induction of SBD-1 expression by S.c.CW. However, TLR4 inhibition only resulted in attenuated SBD-1 mRNA, having no effect on SBD-1 protein expression. Thus, we conclude that S.c.CW can induce SBD-1 expression and that this induction is regulated by the TLR2-MyD88-NF-κB pathway.


Assuntos
Regulação da Expressão Gênica , Probióticos/química , Saccharomyces cerevisiae/química , Ovinos/genética , beta-Defensinas/genética , Ração Animal , Animais , Parede Celular/fisiologia , Células Epiteliais/metabolismo , Rúmen/metabolismo , beta-Defensinas/metabolismo
15.
Acta Odontol Scand ; 78(5): 327-331, 2020 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-31980002

RESUMO

Objective: Type 2 diabetes mellitus (T2DM) is a well-defined risk factor of periodontitis and it can affect expression of human beta-defensins (hBDs) and cathelicidin (LL-37) as well. The aim of the present study was to evaluate the impact of periodontitis and T2DM on salivary concentrations of these antimicrobial peptides.Material and methods: Unstimulated saliva samples, together with full-mouth periodontal recordings were collected from 92 individuals with periodontitis (63 with T2DM and 21 smokers) and 86 periodontally healthy controls (58 with T2DM and 21 smokers). Salivary hBD-1, -2, -3, LL-37, and advanced glycalization end products (AGE) concentrations were measured by enzyme-linked immunosorbent assay.Results: Among the periodontitis patients, T2DM group demonstrated lower levels of hBD-1 (p = .006), hBD-2 (p < .001) and hBD-3 (p < .001), and higher levels of LL-37 (p < .001) compared to systemically healthy controls. When only periodontally healthy controls were included into the analysis, higher hBD-1 (p = .002) and LL-37 (p < .001) levels were found in T2DM patients in comparison to systemically healthy controls. Salivary LL-37 levels were associated with HbA1c and periodontitis, while hBD-2, hBD-3 and levels associated only with HbA1c.Conclusion: In the limits of this study, hyperglycaemia can be proposed as a regulator of salivary hBD and cathelicidin levels. Periodontitis, on the other hand, affects only salivary LL-37 levels.


Assuntos
Peptídeos Catiônicos Antimicrobianos/metabolismo , Diabetes Mellitus Tipo 2/sangue , Periodontite/sangue , Saliva/química , beta-Defensinas/sangue , Adulto , Peptídeos Catiônicos Antimicrobianos/sangue , Estudos de Casos e Controles , Diabetes Mellitus Tipo 2/complicações , Hemoglobina A Glicada , Humanos , Pessoa de Meia-Idade , Periodontite/microbiologia , Periodontite/terapia , Saliva/metabolismo , beta-Defensinas/metabolismo
16.
Dev Comp Immunol ; 103: 103511, 2020 02.
Artigo em Inglês | MEDLINE | ID: mdl-31580833

RESUMO

ß-defensin is a cationic host defense peptide actively participating in host innate immune response against pathogens. In teleost fish, ß-defensin exhibits a diversity in genotypes and functions. Herein, a ß-defensin homolog (PaBD) was identified from ayu, Plecoglossus altivelis, showing multiple tissues' upregulation against Vibrio anguillarum challenge. In vivo experiments revealed that intraperitoneal injection of chemically synthesized mature PaBD (mPaBD) increased the survival rate of V. anguillarum-infected ayu, accompanied by reduced bacterial load and decreased tissue mRNA levels of tumor necrosis factor α (PaTNF-α) and interleukin 1ß (PaIL-1ß). However, in vitro, mPaBD showed weak bactericidal activity against V. anguillarum. Interestingly, mPaBD enhanced phagocytosis, intracellular bacterial killing, and respiratory burst of ayu monocytes/macrophages (MO/MΦ). Moreover, it inhibited mRNA levels of PaIL-1ß and PaTNF-α in MO/MФ upon V. anguillarum infection. In conclusion, PaBD protects ayu against V. anguillarum challenge not only through its direct antibacterial ability, but also through its immunomodulation in MO/MΦ.


Assuntos
Doenças dos Peixes/imunologia , Proteínas de Peixes/metabolismo , Osmeriformes/imunologia , Vibrioses/veterinária , Vibrio/fisiologia , beta-Defensinas/metabolismo , Sequência de Aminoácidos , Animais , Antibacterianos/administração & dosagem , Antibacterianos/farmacologia , Carga Bacteriana/efeitos dos fármacos , Citocinas/genética , Doenças dos Peixes/microbiologia , Doenças dos Peixes/prevenção & controle , Proteínas de Peixes/administração & dosagem , Proteínas de Peixes/genética , Imunomodulação , Macrófagos/imunologia , Macrófagos/microbiologia , Monócitos/imunologia , Monócitos/microbiologia , Osmeriformes/classificação , Osmeriformes/genética , Fagocitose , Filogenia , Explosão Respiratória , Alinhamento de Sequência , Taxa de Sobrevida , Distribuição Tecidual , Vibrio/efeitos dos fármacos , Vibrioses/imunologia , Vibrioses/microbiologia , Vibrioses/prevenção & controle , beta-Defensinas/administração & dosagem , beta-Defensinas/genética
17.
Cell Tissue Res ; 379(3): 601-612, 2020 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-31691005

RESUMO

Differential expression of a variety of proteins in the four major regions of the epididymis contributes to maturation of spermatozoa and region-specific cellular functions as well. Proliferation of epithelial cells of the epididymis is highly controlled and thus is one of the major reasons for the nonoccurrence of cancers in this organ system. The molecular mechanisms and the contribution of region-specific genes in epithelial cell proliferation are not yet fully understood. In this study, for the first time, we analyzed the role of sperm-associated antigen 11a (Spag11a), a caput-specific beta-defensin-like antimicrobial gene in governing epididymal cell proliferation and global gene expression. siRNA-mediated knockdown of Spag11a mRNA in epididymal primary epithelial cells resulted in increased cell proliferation. Out of the 68,842 genes analyzed, 4182 genes were differentially expressed (2154 upregulated and 2028 downregulated). A variety of genes that participate in different cellular processes and pathways were differentially regulated. Genes that are important for epithelial cell proliferation were found to be differentially regulated and these changes were confirmed by real-time PCR. Overexpression of Spag11a in immortalized rat caput epididymal cells resulted in decreased proliferation capacity. Results of this study indicate that Spag11a plays a crucial role in governing epididymal epithelial cell proliferation.


Assuntos
Epididimo/fisiologia , Células Epiteliais/metabolismo , Animais , Western Blotting , Proliferação de Células/fisiologia , Epididimo/citologia , Epididimo/metabolismo , Perfilação da Expressão Gênica , Técnicas de Silenciamento de Genes , Lipídeos/administração & dosagem , Masculino , RNA Mensageiro/antagonistas & inibidores , RNA Mensageiro/biossíntese , RNA Mensageiro/genética , RNA Mensageiro/metabolismo , RNA Interferente Pequeno/administração & dosagem , RNA Interferente Pequeno/genética , Ratos , Ratos Wistar , beta-Defensinas/genética , beta-Defensinas/metabolismo
18.
Am J Physiol Renal Physiol ; 318(2): F468-F474, 2020 02 01.
Artigo em Inglês | MEDLINE | ID: mdl-31841391

RESUMO

Acute pyelonephritis is frequently associated with metabolic acidosis. We previously reported that metabolic acidosis stimulates expression of hypoxia-inducible factor (HIF)-1α-induced target genes such as stromal derived factor-1 and cathelicidin, an antimicrobial peptide. Since the collecting duct (CD) plays a pivotal role in regulating acid-base homeostasis and is the first nephron segment encountered by an ascending microbial infection, we examined the contribution of HIF-1α to innate immune responses elicited by acid loading of an M-1 immortalized mouse CD cell line. Acid loading of confluent M-1 cells was achieved by culture in pH 6.8 medium supplemented with 5-(N-ethyl-N-isopropyl)-amiloride to block Na+/H+ exchange activity for 24 h. Acid loading induced antimicrobial peptide [cathelicidin and ß-defensin (Defb2 and Defb26)] mRNA expression and M-1 cell resistance to uropathogenic Escherichia coli infection to an extent similar to that obtained by inhibition of HIF prolyl hydroxylases, which promote HIF-1α protein degradation. The effect of acid loading on M-1 cell resistance to uropathogenic E. coli infection was reduced by inhibition of HIF-1α (PX-478), and, in combination with prolyl hydroxylase inhibitors, acidosis did not confer additional resistance. Thus, metabolic stress of acidosis triggers HIF-1α-dependent innate immune responses in CD (M-1) cells. Whether pharmacological stabilization of HIF prevents or ameliorates pyelonephritis in vivo warrants further investigation.


Assuntos
Acidose/metabolismo , Peptídeos Catiônicos Antimicrobianos/metabolismo , Infecções por Escherichia coli/prevenção & controle , Subunidade alfa do Fator 1 Induzível por Hipóxia/metabolismo , Túbulos Renais Coletores/metabolismo , Infecções Urinárias/prevenção & controle , Escherichia coli Uropatogênica/patogenicidade , Acidose/imunologia , Animais , Peptídeos Catiônicos Antimicrobianos/genética , Linhagem Celular , Infecções por Escherichia coli/imunologia , Infecções por Escherichia coli/metabolismo , Infecções por Escherichia coli/microbiologia , Interações Hospedeiro-Patógeno , Imunidade Inata , Túbulos Renais Coletores/imunologia , Túbulos Renais Coletores/microbiologia , Camundongos , Prolil Hidroxilases/metabolismo , Estabilidade Proteica , Transdução de Sinais , Regulação para Cima , Infecções Urinárias/imunologia , Infecções Urinárias/metabolismo , Infecções Urinárias/microbiologia , Escherichia coli Uropatogênica/imunologia , beta-Defensinas/metabolismo
19.
J Agric Food Chem ; 68(2): 512-522, 2020 Jan 15.
Artigo em Inglês | MEDLINE | ID: mdl-31870150

RESUMO

Host defense peptides (HDPs) are vital mucosal defense effectors of the innate immune response. The expression of HDPs is inducible in epithelial cells by potent endogenous inducers. Herein, our results demonstrate that sodium butyrate (NaB) induces the expression of porcine ß-defensin-3 (pBD3) and porcine epididymis protein 2 splicing variant C (pEP2C) in a dose- and time-dependent manner, without modifying the production of proinflammatory cytokines, in porcine intestinal epithelial cells (IPEC J2). Moreover, NaB promotes toll-like receptor 2 (TLR2) expression. TLR2 silencing inhibits the pBD3 and pEP2C expression induced by NaB but does not abolish the histone deacetylase (HDAC) inhibitory activity of NaB. We found that NaB activated the nuclear factor-κB (NF-κB) pathway. Importantly, the degree of cell confluence governs the regulatory responses but does not affect the HDAC activity of NaB. Furthermore, epidermal growth factor receptor (EGFR), but not the mitogen-activated protein kinase (MAPK) pathway, is vital during the NaB-induced pBD3 and pEP2C regulation process. We also demonstrated that pBD3 overexpression increases interleukin-18 levels. This study showed that NaB simultaneously induces pBD3 and pEP2C via TLR2 and EGFR in IPEC J2 cells without increasing the risk of a harmful inflammatory response.


Assuntos
Ácido Butírico/farmacologia , Receptores ErbB/metabolismo , Inibidores de Histona Desacetilases/farmacologia , Histona Desacetilases/metabolismo , Receptor 2 Toll-Like/metabolismo , beta-Defensinas/metabolismo , Animais , Células Epiteliais/efeitos dos fármacos , Células Epiteliais/metabolismo , Receptores ErbB/genética , Regulação da Expressão Gênica/efeitos dos fármacos , Histona Desacetilases/genética , Interleucina-18/genética , Interleucina-18/metabolismo , NF-kappa B/genética , NF-kappa B/metabolismo , Suínos , Receptor 2 Toll-Like/genética , beta-Defensinas/genética
20.
Cell Tissue Res ; 379(2): 337-348, 2020 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-31410630

RESUMO

Digital dermatitis (DD), a common ulcerative disease of the bovine foot causing lameness and reducing productivity and animal welfare, is associated with infection by spirochete Treponema bacteria. Topical tetracycline, the most common treatment, has inconsistent cure rates; therefore, new therapeutic options are needed. We compared effects of topical oxytetracycline and vitamin D3 on innate immunity in DD-affected skin. Cows with active DD lesions were treated topically with oxytetracycline or vitamin D3 and skin biopsies were collected from lesions. Tissue samples were examined histologically, transcriptional expression of pro-inflammatory cytokines, Toll-like receptors (TLRs), and host defense peptides assessed, and the presence of specific treponeme species determined. Effects of treatments at a mechanistic level were studied in a human keratinocyte model of treponeme infection. Oxytetracycline promoted hyperplastic scab formation in ulcerated DD lesions and decreased transcriptional expression of Cxcl-8 (neutrophil chemoattractant). Oxytetracycline also reduced numbers of Treponema phagedenis and T. pedis and enhanced Tlr2 mRNA expression. Vitamin D3 did not modify expression of cytokines or Tlrs, or bacterial loads, but enhanced transcription of tracheal antimicrobial peptide (Tap), a key bovine ß-defensin. Combing oxytetracycline and vitamin D3 provides complementary clinical benefits in controlling DD through a combination of antimicrobial, immunomodulatory, and pro-healing activities.


Assuntos
Colecalciferol/uso terapêutico , Dermatite Digital/tratamento farmacológico , Dermatite Digital/microbiologia , Inflamação/tratamento farmacológico , Oxitetraciclina/uso terapêutico , Treponema/fisiologia , beta-Defensinas/genética , Animais , Bovinos , Linhagem Celular , Fatores Quimiotáticos/metabolismo , Colecalciferol/farmacologia , Dermatite Digital/genética , Células Epiteliais/metabolismo , Humanos , Interleucina-8/genética , Interleucina-8/metabolismo , Pele/patologia , Receptor 2 Toll-Like/genética , Receptor 2 Toll-Like/metabolismo , Transcrição Genética , beta-Defensinas/metabolismo
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