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1.
Life Sci ; 254: 117795, 2020 Aug 01.
Artigo em Inglês | MEDLINE | ID: mdl-32417373

RESUMO

AIMS: The primary focus of this study was to explore the effects of cyclic AMP response element-binding protein H (CREBH) on the development of nonalcoholic fatty liver disease (NAFLD). MATERIALS AND METHODS: CREBH knockout (KO) and wildtype (WT) mice were averagely divided into a methionine and choline-deficient (MCD) or high fat (HF) diet group and respective chow diet (CD) groups. Mice were sacrificed after 4-week treatment for MCD model and 24-week treatment for HF model. KEY FINDINGS: Characteristics of nonalcoholic steatohepatitis-related liver fibrosis in KO-MCD/HF group were verified by hepatic histological analyses. Compared with WT-MCD/HF group, levels of plasma ALT and hepatic hydroxyproline increased in KO-MCD/HF group. Significantly higher levels of MCP-1, αSMA, Desmin, COL-1, TIMP-1, TGF-ß1, TGF-ß2 were found while MMP-9 and FGF21 mRNA levels decreased in KO-MCD/HF group. There was also a distinct difference of mRNA levels of TNFα, CTGF and CCND1 in KO-HF group compared with controls. Protein levels of MCP-1, BAX, αSMA, COL-1, TGF-ß1 and SMAD2/3 significantly increased in KO-MCD/HF group and CCND1 was also upregulated in KO-HF group compared to their counterparts. SIGNIFICANCE: CREBH knockout may primarily regulate the TGF-ß1 signaling pathway via TGF-ß2 and FGF21 resulting in more severe inflammation and fibrosis in NAFLD.


Assuntos
Proteína de Ligação ao Elemento de Resposta ao AMP Cíclico/genética , Cirrose Hepática/genética , Hepatopatia Gordurosa não Alcoólica/genética , Fator de Crescimento Transformador beta/metabolismo , Alanina Transaminase/sangue , Animais , Deficiência de Colina , Proteína de Ligação ao Elemento de Resposta ao AMP Cíclico/deficiência , Dieta Hiperlipídica , Fatores de Crescimento de Fibroblastos/biossíntese , Hidroxiprolina/metabolismo , Lipídeos/sangue , Cirrose Hepática/sangue , Cirrose Hepática/metabolismo , Cirrose Hepática/patologia , Masculino , Metionina/deficiência , Camundongos , Camundongos Knockout , Hepatopatia Gordurosa não Alcoólica/sangue , Hepatopatia Gordurosa não Alcoólica/metabolismo , Hepatopatia Gordurosa não Alcoólica/patologia , Transdução de Sinais/genética
2.
Life Sci ; 240: 117096, 2020 Jan 01.
Artigo em Inglês | MEDLINE | ID: mdl-31760097

RESUMO

Aim Liver fibrosis represents a massive global health burden with limited therapeutic options. Thus, the need for curative options is evident. Thus, this study aimed to assess the potential antifibrotic effect of honokiol in Concanavalin A (Con A) induced immunological model of liver fibrosis as well the possible underlying molecular mechanisms. METHODS: Male Sprague-Dawley rats were treated with either Con A (20 mg/kg, IV) and/or honokiol (10 mg/kg, orally) for 4 weeks. Hepatotoxicity indices were as well as histopathological evaluation was done. Hepatic fibrosis was assessed by measuring alpha smooth muscle actin (α-SMA) expression and collagen fibers deposition by Masson's trichrome stain and hydroxyproline content. To elucidate the underlying molecular mechanisms, the effect of honokiol on oxidative stress, inflammatory markers as well as transforming growth factor beta (TGF-ß)/SMAD and mitogen-activated protein kinase (MAPK) pathways was assessed. KEY FINDINGS: Honokiol effectively reversed the hepatotoxicity indices elevations and abnormal histopathological changes induced by Con A. Besides, honokiol attenuated Con A-induced liver fibrosis by down-regulation of hydroxyproline levels, α-SMA expression together with a marked decrease in collagen fibers deposition. Mechanistically Con A induced oxidative stress, provocation of inflammatory responses and activation of TGF-ß/SMAD/MAPK pathways. Contrariwise, honokiol co-treatment significantly restored antioxidant defence mechanisms, down-regulated inflammatory cascades and inhibited TGF-ß/SMAD/MAPK signaling pathways. CONCLUSION: The results provide an evidence for the promising antifibrotic effect of honokiol that could be partially due to suppressing oxidative stress and inflammatory processes as well as inhibition of TGF-ß/SMAD/MAPK signaling pathways.


Assuntos
Compostos de Bifenilo/uso terapêutico , Lignanas/uso terapêutico , Cirrose Hepática/prevenção & controle , Proteínas Quinases Ativadas por Mitógeno/efeitos dos fármacos , Transdução de Sinais/efeitos dos fármacos , Proteínas Smad/efeitos dos fármacos , Fator de Crescimento Transformador beta/efeitos dos fármacos , Actinas/metabolismo , Animais , Concanavalina A , Hidroxiprolina/metabolismo , Inflamação/tratamento farmacológico , Inflamação/patologia , Cirrose Hepática/induzido quimicamente , Cirrose Hepática/patologia , Masculino , Estresse Oxidativo/efeitos dos fármacos , Ratos , Ratos Sprague-Dawley , Análise de Sobrevida
3.
Life Sci ; 242: 117175, 2020 Feb 01.
Artigo em Inglês | MEDLINE | ID: mdl-31843528

RESUMO

AIMS: Ursodeoxycholic acid (UDCA) has been widely used in the treatment of primary biliary cholangitis (PBC) with chronic liver fibrosis, but its detailed mechanism remains unclear. This study was aimed to determine whether autophagic signaling is involved in the therapeutic effect of UDCA on liver fibrosis. METHODS: By using hepatic stellate cell (HSC) line LX2 and CCl4-induced fibrotic rat model, autophagy signaling was investigated by western blotting and mRFP-EGFP-LC3 tandem fluorescent tagged plasmid (ptfLC3) transfection technique. Anti-fibrotic profile was determined by western blotting, qRT-PCR, MTT assay, trypan blue, hydroxyproline assay and Masson staining. KEY FINDINGS: TGFß1 treatment decreased P62 accumulation and increased both autophagosomes and autolysosomes in LX2 cells, thereby elevated autophagic flux. Hydroxychloroquine (HCQ), antagonist of autophagy, was found to dramatically inhibit COL1A2 mRNA expression and cell proliferation in a dose-dependent manner. This coincides with the effect of UDCA intervention on collagen aggradation and cell viability. Meanwhile, UDCA inhibited TGFß1-induced autophagy flux. And rapamycin, agonist of autophagy, was found to impair the anti-fibrotic effect of UDCA. Moreover, study in vivo showed that UDCA alone or in combination with HCQ restored the CCl4-induced liver fibrosis in rodent models with autophagy inhibited profile. SIGNIFICANCE: Taken together, our study revealed that UDCA displays anti-fibrotic role by protecting HSC against production of collagen and inhibiting cellular viability involving autophagy inhibition and provide a new insight into the pharmacological basis of UDCA treatment for hepatic fibrosis.


Assuntos
Autofagia/efeitos dos fármacos , Cirrose Hepática/tratamento farmacológico , Ácido Ursodesoxicólico/uso terapêutico , Animais , Western Blotting , Linhagem Celular , Modelos Animais de Doenças , Humanos , Hidroxicloroquina/farmacologia , Hidroxiprolina/metabolismo , Masculino , Ratos , Ratos Sprague-Dawley , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Fator de Crescimento Transformador beta1/farmacologia
4.
Eur J Pharm Sci ; 141: 105042, 2020 Jan 01.
Artigo em Inglês | MEDLINE | ID: mdl-31634554

RESUMO

Skin aging affects personal image and health. α - lipoic acid (ALA), with excellent free radical scavenging capacity, was used in this research to prepare W/O emulsion. Considering the instability of ALA, ionic liquid strategy was adopted to heighten the solubility of ALA for dissolving in water phase. The mechanism of different ionic liquids (ILs) on skin retention of ALA was investigated by in vitro skin permeation experiment, emulsion quality characterization, rheological test, ATR - FTIR and molecular simulation. The results showed that ionic liquid strategy had a positive influence on the solubilization of ALA. Different ILs were different in skin retention and regulated by skin layers rather than drug release, in which ALA - triethanolamine (ALA - TEOA) presented the best affinity with both stratum corneum (SC) and viable epidermis and dermis (VED), while ALA - N - (2 - Hydroxyethyl) piperidine (ALA - HEPP) as well as ALA - N - (2 - hydroxyethyl) pyrrolidine (ALA - HEPR) showed affinity with either SC or VED respectively. Finally, the emulsion presented brilliant anti - aging efficacy. This study provided a new method of emulsion research and had great significance for the development of topical formulations.


Assuntos
Líquidos Iônicos/administração & dosagem , Envelhecimento da Pele/efeitos dos fármacos , Ácido Tióctico/administração & dosagem , Administração Cutânea , Animais , Emulsões , Hidroxiprolina/metabolismo , Líquidos Iônicos/química , Masculino , Ratos Wistar , Pele/efeitos dos fármacos , Pele/metabolismo , Pele/efeitos da radiação , Absorção Cutânea/efeitos dos fármacos , Envelhecimento da Pele/efeitos da radiação , Ácido Tióctico/química , Raios Ultravioleta/efeitos adversos
5.
Environ Sci Pollut Res Int ; 26(36): 36468-36477, 2019 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-31732951

RESUMO

Paraquat (PQ) induces pulmonary fibrosis, a progressive lung disorder resulting in severe respiratory failure and death. Increased oxidative stress, inflammatory reactions, and multiple fibrotic lesions are major features of PQ-induced lung injury. Diosmin (Dio) is a safe drug that is available for clinical use for vascular disorders. Dio exhibits antioxidant, anti-inflammatory, and antifibrotic activities. Accordingly, the aim of this study was to evaluate the protective effect of diosmin on PQ-induced lung injury in mice and the underlying mechanisms involved. Lung injury was induced by PQ (30 mg/kg, intraperitoneally) in NMRI albino mice and Dio (50 and 100 mg/kg, gavage) was administrated 3 days before PQ and continued for 10 or 24 days. After euthanizing the mice, the biochemical and histopathological markers of lung tissue were determined. PQ significantly increased oxidative stress, inflammatory, and fibrotic markers. PQ increased the level of malonedaldehyde (MDA) and hydroxyproline (HYP) and decreased the level of glutathione (GSH) and catalase activity in the lung. Dio (50 and 100 mg/kg) significantly increased GSH levels and catalase activity and decreased HYP content and MDA levels. In addition, Dio reduced histopathological injuries in hematoxylin and eosin-stained and Masson's trichrome-stained sections. These findings suggest that Dio has protective effects against PQ-induced lung injury, which may be due to its antioxidant, anti-inflammatory, and antifibrotic effects.


Assuntos
Antioxidantes/farmacologia , Diosmina/farmacologia , Lesão Pulmonar/prevenção & controle , Estresse Oxidativo/efeitos dos fármacos , Paraquat/toxicidade , Fibrose Pulmonar/prevenção & controle , Animais , Fibrose , Glutationa/metabolismo , Hidroxiprolina/metabolismo , Lesão Pulmonar/induzido quimicamente , Lesão Pulmonar/metabolismo , Lesão Pulmonar/patologia , Masculino , Camundongos , Camundongos Endogâmicos , Fibrose Pulmonar/induzido quimicamente , Fibrose Pulmonar/metabolismo , Fibrose Pulmonar/patologia
6.
Neurotoxicol Teratol ; 76: 106836, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-31593814

RESUMO

Pesticides despite being agents that protect the plants and humans from noxious pests, are infamous for their potential to cause detrimental health issues in nontargeted species. In order to ascertain the latter, a set of experiments were conducted by exposing early chick embryos to a widely used combination insecticide (Ci, 50% chlorpyrifos and 5% cypermethrin). The results revealed a myriad of congenital defects pertaining to craniofacial development such as anophthalmia, microphthalmia, exencephaly as well as deformed beak and cranial structures. These teratological manifestations could be attributed to the Ci induced alteration in the titre of major regulators of neurulation and ossification. Therefore, the mRNA and/or the protein level expression pattern of genes which are reported to be involved in the craniofacial development were studied at selected time points of embryonic development. The analysis of the result showed that there have been significant alternations in the expression patterns of the signalling molecules such as SHH, WNTs, CDH1, CDH2, L1CAM, PAX6, HOX, PCNA, GLI3, BMP7, FGF8, GLIs, SOX9, RUNX2, DLX5, COL10A1, CASPASE3 etc. on embryonic days 2, 4 and/or 10. Concurrently, on day 10, whole-mount skeletal staining and biochemical estimation of hydroxyproline were carried out in the cranial tissues of the embryos. The overall result of the current study indicates that exposure to Ci during early development impede the crucial regulatory signals that orchestrate the morphogenesis of cranial neural crest cells thereby hindering the normal progression of neural tube and endochondral ossification which collectively lead to craniofacial dysmorphism in domestic chicks.


Assuntos
Anormalidades Craniofaciais/induzido quimicamente , Inseticidas/toxicidade , Transdução de Sinais/efeitos dos fármacos , Transdução de Sinais/genética , Animais , Bico/anormalidades , Química Encefálica/efeitos dos fármacos , Embrião de Galinha , Galinhas , Clorpirifos/toxicidade , Anormalidades Craniofaciais/mortalidade , Anormalidades Craniofaciais/fisiopatologia , Feminino , Regulação da Expressão Gênica no Desenvolvimento/efeitos dos fármacos , Hidroxiprolina/metabolismo , Gravidez , Efeitos Tardios da Exposição Pré-Natal , Piretrinas/toxicidade , RNA Mensageiro/biossíntese , RNA Mensageiro/genética
7.
Phys Chem Chem Phys ; 21(42): 23338-23345, 2019 Nov 14.
Artigo em Inglês | MEDLINE | ID: mdl-31617504

RESUMO

Nature exploits different strategies for enhancing the catalytic activity of enzymes, often resorting to producing beneficial mutations. The case of post-translational proline hydroxylation mutation in the active site of polysaccharide deacetylase (PDA) Bc1960 from Bacillus cereus is an interesting example of how small chemical modifications can cause significant improvements in enzymatic activity. In the present study the deacetylation mechanism promoted by both OH-proline (2Hyp) and standard proline (Pro) containing PDA is investigated using density functional theory. Although the mechanism presented for the two examined enzymes is in agreement with protease catalysis in metalloenzymes, the analysis along the potential energy surface (PES) reveals that the intermediate and product benefit energetically from the presence of the hydroxyl group on the proline. Our calculations provide evidence that for PDA-2Hyp, the hydrogen bond network established by the -OH group on the Cα of the proline with its closest neighbors stabilizes the transition states and, consequently, the reaction takes advantage of this. These results further contribute towards explaining the different catalytic activity experimentally observed for the polysaccharide deacetylase enzymes.


Assuntos
Amidoidrolases/metabolismo , Hidroxiprolina/metabolismo , Amidoidrolases/química , Bacillus cereus/enzimologia , Proteínas de Bactérias/química , Proteínas de Bactérias/metabolismo , Sítios de Ligação , Biocatálise , Ligação de Hidrogênio , Hidroxiprolina/química , Simulação de Dinâmica Molecular , Estrutura Terciária de Proteína , Termodinâmica
8.
Metabolomics ; 15(9): 123, 2019 09 06.
Artigo em Inglês | MEDLINE | ID: mdl-31493001

RESUMO

INTRODUCTION: German shepherd dogs (GSDs) are a popular breed affected by numerous disorders. Few studies have explored genetic variations that influence canine blood metabolite levels. OBJECTIVES: To investigate genetic variants affecting the natural metabolite variation in GSDs. METHODS: A total of 82 healthy GSDs were genotyped on the Illumina CanineHD Beadchip, assaying 173,650 markers. For each dog, 74 metabolites were measured through liquid and gas chromatography mass spectrometry (LC-MS and GC-MS) and were used as phenotypes for genome-wide association analyses (GWAS). Sliding window and homozygosity analyses were conducted to fine-map regions of interest, and to identify haplotypes and gene dosage effects. RESULTS: Summary statistics for 74 metabolites in this population of GSDs are reported. Forty-one metabolites had significant associations at a false discovery rate of 0.05. Two associations were located around genes which encode for enzymes for the relevant metabolites: 4-hydroxyproline was significantly associated to D-amino acid oxidase (DAO), and threonine to L-threonine 3-dehydrogenase (LOC477365). Three of the top ten haplotypes associated to 4-hydroxyproline included at least one SNP on DAO. These haplotypes occurred only in dogs with the highest 15 measurements of 4-hydroxyproline, ranging in frequency from 16.67 to 20%. None of the dogs were homozygous for these haplotypes. The top two haplotypes associated to threonine included SNPs on LOC477365 and were also overrepresented in dogs with the highest 15 measurements of threonine. These haplotypes occurred at a frequency of 90%, with 80% of these dogs homozygous for the haplotypes. In dogs with the lowest 15 measurements of threonine, the haplotypes occurred at a frequency of 26.67% and 0% homozygosity. CONCLUSION: DAO and LOC477365 were identified as candidate genes affecting the natural plasma concentration of 4-hydroxyproline and threonine, respectively. Further investigations are needed to validate the effects of the variants on these genes.


Assuntos
Cães/genética , Metaboloma , Polimorfismo de Nucleotídeo Único , Oxirredutases do Álcool/genética , Animais , D-Aminoácido Oxidase/genética , Cães/sangue , Feminino , Cromatografia Gasosa-Espectrometria de Massas/métodos , Estudo de Associação Genômica Ampla/métodos , Haplótipos , Hidroxiprolina/metabolismo , Masculino
9.
Pak J Pharm Sci ; 32(3 Special): 1387-1393, 2019 May.
Artigo em Inglês | MEDLINE | ID: mdl-31551220

RESUMO

The histological and ultrastructural changes of skin of Wistar rats were compared with 10600nm CO2 dot laser irradiation and external use of A acid preparation. The effects of dot matrix CO2 laser and Nd: YAG laser (quasi long pulse width 10600nm wavelength) on the expression of matrix metalloproteinase -1 (MMP-1) and matrix metalloproteinase inhibitor -1 (TIMP-1) in skin of natural aging mice were investigated, and the skin tender mechanism was further explored. To explore the long-term efficacy and timeliness of 10600nm CO2 dot laser and A acid treatment, so as to provide reference for clinical practice. The skin of hair on the back of mice was irradiated with dot CO2 laser and Nd: YAG laser. They were taken at 2 weeks, 1 month and 3 months after irradiation respectively. The dynamic changes of skin HSP47, HSP70 and TGF - ß 1 were detected by immunohistochemistry, and the difference of two laser irradiation effects was compared. 45 female Wistar rats were randomly divided into 9 groups. The back skin was used as the experimental observation area. After the depilation, the observation was divided into four parts by using the cross shaped marking line: the left side of the proximal end was the normal control group (A area). At the proximal end, the left side was the vitamin A acid group (B area). The right side of the tail was the combined treatment group (C area), and the right side of the proximal end was the dot laser group (D area). The C and D regions irradiated 10600 CO2 dot matrix laser 1 times at the beginning of the experiment. The parameters were: energy 15mJ, energy density 5%, frequency 300Hz. The fig. is square, 10mm * 10mm; after the laser irradiation, the B and C areas begin to wipe the 0.025% dimensional A cream every day for 3 weeks. The positive expression of MMP-1 and TIMP-1 in dot CO2 laser and Nd: YAG laser irradiation area was most obvious at 2 weeks. The MMP-1 gray value (115.14 + 5.23) and TIMP-1 gray value (104.01 + 3.15) of the dot matrix laser irradiated area were lower than the MMP-1 gray value (121.75 + 4.39) and TIMP-1 gray value (109.26 + 3.88) in the Nd: YAG laser irradiation area. That is, the positive expression of CO2 laser irradiation area was higher, and the difference was statistically significant (P < 0.05).


Assuntos
Hidroxiprolina/metabolismo , Lasers de Gás/uso terapêutico , Pele/efeitos da radiação , Envelhecimento , Animais , Feminino , Proteínas de Choque Térmico HSP47/metabolismo , Lasers de Estado Sólido , Metaloproteinase 13 da Matriz/metabolismo , Camundongos , Pele/metabolismo , Pele/ultraestrutura , Inibidor Tecidual de Metaloproteinase-1/metabolismo
10.
Molecules ; 24(18)2019 Sep 14.
Artigo em Inglês | MEDLINE | ID: mdl-31540139

RESUMO

Thymus sipyleus Boiss. subsp. rosulans (Borbas) Jalas (TS) is a commonly used plant in the treatment of various complaints, including skin wounds in Turkish folk medicine. Despite the widespread traditional use of TS, there is not any scientific report confirming the effectiveness of this plant on the healing process. This research aimed to investigate the effects of different extracts obtained from TS on biological events during wound healing, on a cellular basis. In this context, proliferative activities of the extracts, as well as the effects on wound closure and hydroxyproline synthesis, were determined. In addition to wound healing properties, the antioxidant, antibacterial and anti-inflammatory activities of the extracts were evaluated. Decoction (D) and infusion (I) extracts contained the highest amount of phenolic content and showed the most potent activity against DPPH radical. All extracts exhibited complete protection against the damage induced by hydrogen peroxide (H2O2) by increasing cell viability compared to only H2O2-treated groups, both in co-treatment and pre-treatment protocols. None of the extracts exhibited cytotoxic activity, and most of the extracts from the TS stimulated fibroblast proliferation and migration. All TS extracts exert anti-inflammatory activity by suppressing the overproduction of tumor necrosis factor-alpha (TNF-α) and nitric oxide (NO). The most pronounced activity on hydroxyproline synthesis was observed in D extract. In summary, it was observed that TS extracts can promote the healing process by enhancing fibroblast migration, proliferation and collagen synthesis as well as suppressing pro-inflammatory cytokines. The obtained data in this work support the traditional use of TS as a valuable plant-based compound for the treatment of wounds.


Assuntos
Anti-Infecciosos/farmacologia , Anti-Inflamatórios/farmacologia , Antioxidantes/farmacologia , Fibroblastos/metabolismo , Extratos Vegetais/farmacologia , Thymus (Planta)/química , Cicatrização/efeitos dos fármacos , Células 3T3 , Animais , Anti-Infecciosos/química , Anti-Inflamatórios/química , Antioxidantes/química , Movimento Celular/efeitos dos fármacos , Avaliação Pré-Clínica de Medicamentos , Fibroblastos/patologia , Hidroxiprolina/metabolismo , Camundongos , Óxido Nítrico/metabolismo , Extratos Vegetais/química , Fator de Necrose Tumoral alfa/metabolismo
11.
Adv Gerontol ; 32(3): 331-337, 2019.
Artigo em Russo | MEDLINE | ID: mdl-31512418

RESUMO

Aging of extracellular proteins colloidal systems is one of major synchronizing mechanism in mammal`s «biological clock¼. We hypothesized that induced controllable modification of connective tissue composition could reverse aging. In murine experimental models collagenase was used for selective destruction of old collagen. Oxygen consumption, urine hydroxyproline excretion, density and distribution of mature and old collagen and elastine fibers in dermal biopsies were determined. Collagenase injections significantly increased hydroxyproline excretion. We observed reduced density of mature and old collagen fibers and increased oxygen consumption in dermal biopsies after course of collagenase injections. Collagenase treatment intensified the destruction of mature and old collagen matrix and enhanced synthesis of new collagen and elastine fibers. Furthermore oxygen consumption increased. Our findings can be considered as indicator of collagenase systemic anti-aging (rejuvenation) activity.


Assuntos
Envelhecimento , Colágeno , Envelhecimento/efeitos dos fármacos , Animais , Colágeno/metabolismo , Colagenases/farmacologia , Hidroxiprolina/metabolismo , Camundongos , Modelos Animais , Consumo de Oxigênio/efeitos dos fármacos , Pele/efeitos dos fármacos
12.
Int Immunopharmacol ; 75: 105755, 2019 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-31377591

RESUMO

Pulmonary fibrosis is an irreversible lung disorder with predictable decline in lung function leading to respiratory insufficiency. Incidence of pulmonary fibrosis has been apparently increasing worldwide. Though aetiology of this disease remains unclear, potential roles of infection, disordered cell biology, genetic influence etc. have been proposed. Pirfenidone and nintedanib are the only two US FDA approved drugs to treat pulmonary fibrosis. Autophagy is a catabolic intracellular pathway that plays a crucial role in maintaining cellular homeostasis, which is involved in many disorders including fibrotic diseases. The present study investigated the role of Nimbolide, an important active constituent of Neem in TGF-ß1 induced in vitro and bleomycin induced in vivo model of pulmonary fibrosis, with a slight emphasis on regulation of fibrosis related autophagy. Protein expression studies showed significant reduction in mesenchymal, fibrotic markers and a substantial up regulation of epithelial markers upon treatment with Nimbolide. Nimbolide regulated autophagy signaling by dampening LC-3 and p-62 expression and increasing Beclin 1 expression as evidenced by immunohistochemistry and confocal microscopy. Our study demonstrates Nimbolide as a potent anti-fibrotic agent and its ability to regulate fibrosis associated autophagy.


Assuntos
Limoninas/uso terapêutico , Fibrose Pulmonar/tratamento farmacológico , Animais , Autofagia/efeitos dos fármacos , Bleomicina , Líquido da Lavagem Broncoalveolar/química , Linhagem Celular , Movimento Celular/efeitos dos fármacos , Sobrevivência Celular/efeitos dos fármacos , Transição Epitelial-Mesenquimal/efeitos dos fármacos , Humanos , Hidroxiprolina/metabolismo , L-Lactato Desidrogenase/metabolismo , Limoninas/farmacologia , Masculino , Camundongos , Óxido Nítrico/metabolismo , Fibrose Pulmonar/induzido quimicamente , Fibrose Pulmonar/patologia , Fator de Crescimento Transformador beta1
13.
Int Wound J ; 16(6): 1426-1432, 2019 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-31448554

RESUMO

Hyperthermic intraperitoneal chemotherapy (HIPEC) has cytotoxic effects on tumour cells but also negative impacts on anastomotic healing. Platelet-rich-plasma (PRP) is used for wound care but data about effects on gastrointestinal anastomosis are limited. In this experimental study, we aimed to investigate the effects of PRP application on colon anastomosis in rats those received HIPEC with cisplatin. Five rats were sacrificed to obtain PRP gel. Thirty rats were divided into three groups; Group 1: control group, Group 2: colon anastomosis and HIPEC with cisplatin, and Group 3: colon anastomosis enhanced by PRP and HIPEC with cisplatin. The rats were re-operated on postoperative day seven and anastomotic bursting pressure (ABP) was recorded. Also, tissue samples were taken for hydroxyproline assessment and histopathological examination. There were significant differences in ABP between Groups 2 and 3, and also those groups had lower ABP compared with the control group. Group 3 had significantly higher hydroxyproline levels and had better histopathological findings than group 2. According to our findings, we suggest that PRP application improves the anastomotic healing by increasing anastomotic bursting pressure, hydroxyproline levels, and decreasing inflammatory response. Further clinical studies are needed to prove our hypothesis.


Assuntos
Anastomose Cirúrgica , Antineoplásicos/administração & dosagem , Colo/cirurgia , Hipertermia Induzida , Plasma Rico em Plaquetas , Cicatrização , Animais , Cisplatino/administração & dosagem , Colo/metabolismo , Edema/patologia , Géis , Hidroxiprolina/metabolismo , Linfócitos/metabolismo , Macrófagos/metabolismo , Modelos Animais , Neutrófilos/metabolismo , Ratos Wistar
14.
Lung ; 197(5): 541-549, 2019 10.
Artigo em Inglês | MEDLINE | ID: mdl-31392398

RESUMO

PURPOSE: Growth hormone-releasing hormone (GHRH) is a 44-amino acid peptide that regulates growth hormone (GH) secretion. We hypothesized that a GHRH receptor (GHRH-R) antagonist, MIA-602, would inhibit bleomycin-induced lung inflammation and/or fibrosis in C57Bl/6J mice. METHODS: We tested whether MIA-602 (5 µg or vehicle given subcutaneously [SC] on days 1-21) would decrease lung inflammation (at day 14) and/or fibrosis (at day 28) in mice treated with intraperitoneal (IP) bleomycin (0.8 units on days 1, 3, 7, 10, 14, and 21). Bleomycin resulted in inflammation and fibrosis around airways and vessels evident histologically at days 14 and 28. RESULTS: Inflammation (histopathologic scores assessed blindly) was visibly less evident in mice treated with MIA-602 for 14 days. After 28 days, lung hydroxyproline (HP) content increased significantly in mice treated with vehicle; in contrast, lung HP did not increase significantly compared to naïve controls in mice treated with GHRH-R antagonist. GHRH-R antagonist increased basal and maximal oxygen consumption of cultured lung fibroblasts. Multiple genes related to chemotaxis, IL-1, chemokines, regulation of inflammation, and extracellular signal-regulated kinases (ERK) were upregulated in lungs of mice treated with bleomycin and MIA-602. MIA-602 also prominently suppressed multiple genes related to the cellular immune response including those for T-cell differentiation, receptor signaling, activation, and cytokine production. CONCLUSIONS: MIA-602 reduced lung inflammation and fibrosis due to bleomycin. Multiple genes related to immune response and T-cell functions were downregulated, supporting the view that MIA-602 can modulate the cellular immune response to bleomycin lung injury.


Assuntos
Bleomicina , Hormônio Liberador de Hormônio do Crescimento/antagonistas & inibidores , Antagonistas de Hormônios/farmacologia , Pulmão/efeitos dos fármacos , Pneumonia/prevenção & controle , Fibrose Pulmonar/prevenção & controle , Sermorelina/análogos & derivados , Animais , Células Cultivadas , Citoproteção , Modelos Animais de Doenças , MAP Quinases Reguladas por Sinal Extracelular/metabolismo , Fibroblastos/efeitos dos fármacos , Fibroblastos/metabolismo , Fibroblastos/patologia , Regulação da Expressão Gênica , Hormônio Liberador de Hormônio do Crescimento/metabolismo , Hidroxiprolina/metabolismo , Mediadores da Inflamação/metabolismo , Pulmão/metabolismo , Pulmão/patologia , Masculino , Camundongos Endogâmicos C57BL , Pneumonia/induzido quimicamente , Pneumonia/metabolismo , Pneumonia/patologia , Fibrose Pulmonar/induzido quimicamente , Fibrose Pulmonar/metabolismo , Fibrose Pulmonar/patologia , Sermorelina/farmacologia , Transdução de Sinais
15.
Food Chem ; 301: 125302, 2019 Dec 15.
Artigo em Inglês | MEDLINE | ID: mdl-31387034

RESUMO

The autolysis of sea cucumber is caused by depolymerisation of collagen fibres and unfolding of fibrils. In order to highlight the role of collagenase in sea cucumber autolysis, collagen fibres from sea cucumber were hydrolysed with collagenase type I. Electron microscopy (EM) results indicated the collagenase caused partial depolymerisation of collagen fibres into fibrils due to the fracture of proteoglycan interfibrillar bridges, as well as uncoiling of collagen fibrils. Chemical analysis and SDS-PAGE both indicated collagenase induced a time-dependent release of glycosaminoglycans (GAGs) and soluble proteins, which further demonstrated the degradation of proteoglycan interfibrillar bridges. Collagenase also degraded collagens by releasing soluble hydroxyproline (Hpy), with the dissolution rate of Hyp reaching 11.11% after 72 h. Fourier transform infrared analysis showed that collagenase caused the reduction of intermolecular interactions and structural order of collagen. Hence, collagenase participated in the autolysis of sea cucumber by deteriorating both macromolecular and monomeric collagens.


Assuntos
Colágeno/química , Colagenases/química , Stichopus/química , Animais , Autólise , Colágeno/metabolismo , Colagenases/metabolismo , Eletroforese em Gel de Poliacrilamida , Glicosaminoglicanos/metabolismo , Hidrólise , Hidroxiprolina/metabolismo , Microscopia Eletrônica de Varredura , Microscopia Eletrônica de Transmissão , Proteoglicanas/química , Proteoglicanas/metabolismo , Espectroscopia de Infravermelho com Transformada de Fourier , Stichopus/anatomia & histologia
16.
Invest Ophthalmol Vis Sci ; 60(10): 3422-3431, 2019 08 01.
Artigo em Inglês | MEDLINE | ID: mdl-31390655

RESUMO

Purpose: The degenerative corneal disease keratoconus is a leading indicator for corneal transplant with an unknown etiology. We recently identified the activation of the integrated stress response (ISR) in ex vivo human corneas and in vitro cell culture. Utilizing small molecules to modulate the ISR we sought to investigate the effects of stimulating the ISR in healthy cells to recapitulate aspects of the in vitro keratoconic phenotype and whether relieving the ISR signaling would recover the disease phenotype. Methods: Corneal fibroblasts were extracted from patients undergoing corneal transplant or unaffected cadaverous donor limbal rings. Cells were exposed to the DNA damage-inducible protein (GADD34) inhibitor SAL003 to stimulate the ISR, or Trans-ISRIB to relieve ISR signaling pathway. Collagen production was assessed through hydroxyproline production, Sirius Red incorporation, or quantitative (q)PCR. Western blotting, hydroxyproline, and qPCR were used to assess components of the ISR pathway and collagen production. Results: ISR stimulation through SAL003 resulted in significant decrease of hydroxyproline and COL1A1 transcription and eventual apoptosis in normal fibroblasts. Patient (KC) fibroblast production of hydroxyproline was increased in response to ISRIB, while matrix metalloproteinase (MMP)9 production was lowered. The prospective biomarker of keratoconus prolactin-inducible factor was also upregulated in KC fibroblast cultures in response to ISRIB. Inflammatory markers TNFα and IL-1ß were unaffected. Conclusions: Activation of the ISR is sufficient to recapitulate many key aspects of the KC phenotype in unaffected cells in vitro. Inhibition of the ISR also relieves many of the hallmarks of KC in affected cells. Therefore, targeting of the ISR through small molecules is a potential therapeutic path for small molecule treatment of keratoconus.


Assuntos
Acetamidas/farmacologia , Ceratócitos da Córnea/efeitos dos fármacos , Cicloexilaminas/farmacologia , Inibidores Enzimáticos/farmacologia , Ceratocone/patologia , Estresse Fisiológico/efeitos dos fármacos , Western Blotting , Contagem de Células , Células Cultivadas , Colágeno Tipo I/genética , Ceratócitos da Córnea/metabolismo , Humanos , Hidroxiprolina/metabolismo , Metaloproteinase 2 da Matriz/genética , Metaloproteinase 9 da Matriz/genética , Fenótipo , Estudos Prospectivos , Proteína Fosfatase 1/antagonistas & inibidores , Reação em Cadeia da Polimerase em Tempo Real
17.
Invest Ophthalmol Vis Sci ; 60(8): 2895-2903, 2019 07 01.
Artigo em Inglês | MEDLINE | ID: mdl-31266061

RESUMO

Purpose: The proinflammatory cytokine interleukin (IL)-1 is implicated in corneal ulceration and promotes collagen degradation by corneal fibroblasts cultured in a three-dimensional (3D) collagen gel. Epigallocatechin-3-gallate (EGCG), the principal polyphenol in extracts of green tea, has various beneficial health effects, some of which appear to be mediated through direct or indirect inhibition of protease activity. We therefore examined the effect of EGCG on IL-1ß-induced collagen degradation by corneal fibroblasts embedded in a collagen gel. Methods: Human corneal fibroblasts were cultured in a type I collagen gel. Collagen degradation was assessed by measurement of hydroxyproline in acid hydrolysates of culture supernatants. The expression of urokinase-type plasminogen activator (uPA) was examined by real-time and RT-PCR analysis and by fibrin zymography, and that of the collagenase matrix metalloproteinase 1 (MMP1) was detected by immunoblot analysis. Results: EGCG inhibited IL-1ß-induced, plasminogen-dependent collagen degradation by corneal fibroblasts in a concentration-dependent manner. It also attenuated the IL-1ß-induced expression of uPA at both mRNA and protein levels. EGCG inhibited the IL-1ß-induced conversion of exogenous plasminogen to plasmin as well as the plasminogen-dependent activation of pro-MMP1 in the 3D cultures without a substantial effect on pro-MMP1 abundance. Conclusions: EGCG inhibits IL-1ß-induced collagen degradation by corneal fibroblasts, with this effect likely being mediated by suppression of the upregulation of uPA, the uPA-mediated conversion of plasminogen to plasmin, and the plasmin-mediated activation of pro-MMP1. EGCG thus warrants further investigation as a potential treatment for corneal ulcer.


Assuntos
Antioxidantes/farmacologia , Catequina/análogos & derivados , Colágeno/metabolismo , Ceratócitos da Córnea/efeitos dos fármacos , Interleucina-1beta/antagonistas & inibidores , Ativador de Plasminogênio Tipo Uroquinase/genética , Catequina/farmacologia , Sobrevivência Celular/efeitos dos fármacos , Células Cultivadas , Ceratócitos da Córnea/metabolismo , Relação Dose-Resposta a Droga , Fibrina/metabolismo , Fibrinolisina/metabolismo , Regulação da Expressão Gênica/efeitos dos fármacos , Regulação da Expressão Gênica/fisiologia , Humanos , Hidroxiprolina/metabolismo , Interleucina-1beta/farmacologia , Metaloproteinase 1 da Matriz/metabolismo , NF-kappa B/metabolismo , Reação em Cadeia da Polimerase em Tempo Real
18.
Mol Immunol ; 114: 72-80, 2019 10.
Artigo em Inglês | MEDLINE | ID: mdl-31344551

RESUMO

Scleroderma is an inflammatory autoimmune disease characterized by extensive tissue fibrosis. The imbalance of effector T (Teff) and regulatory T (Treg) cells and the production of autoantibodies contribute to the pathogenesis of this disease. Metformin (MET) has anti-inflammatory and anti-fibrotic effects, but its effect on the in vivo pathogenesis of scleroderma remains unknown. Therefore, we investigated the potential therapeutic effects of MET treatment of mice with bleomycin (BLM)-induced scleroderma. Scleroderma was induced in female C57BL mice by daily subcutaneous injections of BLM for 28 days. After each 2 h BLM injection, mice received MET (200, 100 or 50 mg/kg) or saline (control) by intraperitoneal injection. At the end of the fourth week, spleen mononuclear cells were collected for flow cytometry analysis. Skin samples were harvested for immunohistochemistry and quantification of other biological parameters.Our results showed that BLM increased dermal thickness, collagen deposition, and hydroxyproline level, and MET markedly mitigated these effects. MET also restored the Treg/Teff cell balance. Accordingly, the level of IL-17A and RORγt (related to Th17 cells) decreased, but Foxp3 (related to Treg function) increased in a dose-dependent manner. In addition, MET treatment inhibited spleen germinal center formation. These results indicate that the immunomodulatory and anti-fibrosis effects of MET on BLM-induced scleroderma are mediated by the upregulation of Treg cell differentiation, inhibition of Teff cell differentiation, and suppression of spleen germinal center formation. These results suggest that MET may be a potential therapeutic for scleroderma.


Assuntos
Bleomicina/farmacologia , Centro Germinativo/efeitos dos fármacos , Metformina/farmacologia , Dermatopatias/induzido quimicamente , Dermatopatias/tratamento farmacológico , Baço/efeitos dos fármacos , Linfócitos T Reguladores/efeitos dos fármacos , Animais , Diferenciação Celular/efeitos dos fármacos , Colágeno/metabolismo , Modelos Animais de Doenças , Feminino , Fibrose/tratamento farmacológico , Fibrose/metabolismo , Centro Germinativo/metabolismo , Hidroxiprolina/metabolismo , Interleucina-17/metabolismo , Camundongos , Camundongos Endogâmicos C57BL , Membro 3 do Grupo F da Subfamília 1 de Receptores Nucleares/metabolismo , Pele/efeitos dos fármacos , Pele/metabolismo , Dermatopatias/metabolismo , Baço/metabolismo , Linfócitos T Reguladores/metabolismo , Células Th17/efeitos dos fármacos , Células Th17/metabolismo
19.
Int Urol Nephrol ; 51(7): 1121-1127, 2019 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-31161522

RESUMO

PURPOSE: Dietary hydroxyproline may be involved in the endogenous synthesis of oxalate. Glycolate, produced during the metabolism of hydroxyproline, may exert physicochemical effects on urinary calcium by virtue of its dihydroxycarboxylic acid structure. The aim of this study was to investigate these possible stone-risk scenarios. METHODS: We modelled the effect of different glycolic acid concentrations on ionized calcium (iCa2+) and relative supersaturation (RSS) of calcium oxalate (CaOx) using the program JESS. Thereafter, three healthy white males and two healthy black males ingested 30 g gelatin for 3 days. 24-h urines were collected at baseline and after completion of the protocol. Urines were analysed for physicochemical risk factors and for iCa2+ and glycolic acid. Speciation concentrations and RSS values were calculated. RESULTS: Theoretical modelling showed that binding between calcium and glycolate does not occur and that iCa2+ and RSS CaOx are unaffected. However, after ingestion of hydroxyproline, iCa2+ decreased significantly. Urinary pH and glycolate increased significantly. Oxalate excretion and RSS CaOx did not change CONCLUSIONS: We attribute the decrease in iCa2+ to increases in the concentrations of several Ca-phosphate species, the formation of which is due to the increase in pH. We speculate that the absence of an increase in oxalate excretion despite an increase in glycolate excretion may be due to the mixed racial composition of our test group in which some pathways may be preferred to others. Our findings alert stone researchers to the importance of measuring urinary pH in their workup of subjects and to select racially homogenous groups for investigation.


Assuntos
Oxalato de Cálcio , Glicolatos , Hidroxiprolina/metabolismo , Nefrolitíase , Adulto , Fenômenos Bioquímicos , Oxalato de Cálcio/metabolismo , Oxalato de Cálcio/urina , Exposição Dietética , Glicolatos/metabolismo , Glicolatos/urina , Voluntários Saudáveis , Humanos , Concentração de Íons de Hidrogênio , Masculino , Nefrolitíase/diagnóstico , Nefrolitíase/metabolismo , Medição de Risco/métodos , Fatores de Risco , Urinálise/métodos
20.
J Sci Food Agric ; 99(13): 5752-5759, 2019 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-31162681

RESUMO

BACKGROUND: Sea cucumber (Stichopus japonicus) is easy to autolysis in response to a variety of environmental and mechanical factors. In the current study, collagen fibres were extracted from fresh sea cucumber body wall and then incubated with endogenous matrix metalloprotease (MMP) of sea cucumber. Scanning electron microscopy (SEM), differential scanning calorimetry (DSC), chemical analysis and sodium dodecyl sulphate polyacrylamide gel electrophoresis (SDS-PAGE) analysis were utilized to demonstrate the changes in collagen fibres, collagen fibrils and collagen proteins. Moreover, a verification experiment was also carried out to confirm the contribution of MMP to the autolysis of sea cucumber. RESULTS: Endogenous MMP caused complete depolymerization of collagen fibres into smaller collagen fibril bundles and collagen fibrils due to the fracture of proteoglycan interfibrillar bridges. Meanwhile, endogenous MMP also caused partial degradation of collagen fibrils by releasing soluble hydroxyproline and pyridinium cross-links. Furthermore, the treatment with MMP inhibitor (1,10-phenanthroline) prevented the autolysis of tissue blocks from S. japonicus dermis. CONCLUSION: Endogenous MMP was the key enzyme in the autolysis of sea cucumber, while its action still focused on high-level structures of collagens especially collagen fibres. © 2019 Society of Chemical Industry.


Assuntos
Autólise , Metaloproteases/metabolismo , Stichopus/enzimologia , Stichopus/fisiologia , Animais , Colágeno/metabolismo , Colágeno/ultraestrutura , Hidroxiprolina/metabolismo , Metaloproteases/genética , Microscopia Eletrônica de Varredura , Stichopus/ultraestrutura
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