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1.
Int J Mol Sci ; 22(11)2021 Jun 01.
Artigo em Inglês | MEDLINE | ID: mdl-34206086

RESUMO

Tuberculosis (TB) is an important infectious disease and a public health problem. The organs most frequently affected by TB are the lungs; despite this, it has been reported that TB patients suffer from depression and anxiety, which have been attributed to social factors. In previous experimental work, we observed that the extensive pulmonary inflammation characteristic of TB with high cytokine production induces neuroinflammation, neuronal death and behavioral abnormalities in the absence of brain infection. The objective of the present work was to reduce this neuroinflammation and avoid the psycho-affective disorders showed during pulmonary TB. Glucocorticoids (GCs) are the first-line treatment for neuroinflammation; however, their systemic administration generates various side effects, mostly aggravating pulmonary TB due to immunosuppression of cellular immunity. Intranasal administration is a route that allows drugs to be released directly in the brain through the olfactory nerve, reducing their doses and side effects. In the present work, dexamethasone's (DEX) intranasal administration was evaluated in TB BALB /c mice comparing three different doses (0.05, 0.25 and 2.5 mg/kg BW) on lung disease evolution, neuroinflammation and behavioral alterations. Low doses of dexamethasone significantly decreased neuroinflammation, improving behavioral status without aggravating lung disease.


Assuntos
Encéfalo/efeitos dos fármacos , Dexametasona/farmacologia , Inflamação/tratamento farmacológico , Tuberculose Pulmonar/tratamento farmacológico , Administração Intranasal , Animais , Ansiedade/complicações , Ansiedade/tratamento farmacológico , Ansiedade/patologia , Encéfalo/patologia , Depressão/complicações , Depressão/tratamento farmacológico , Depressão/patologia , Modelos Animais de Doenças , Glucocorticoides/farmacologia , Humanos , Inflamação/complicações , Inflamação/microbiologia , Inflamação/patologia , Camundongos , Mycobacterium tuberculosis/efeitos dos fármacos , Mycobacterium tuberculosis/patogenicidade , Tuberculose Pulmonar/complicações , Tuberculose Pulmonar/microbiologia , Tuberculose Pulmonar/patologia
2.
Int J Mol Sci ; 22(11)2021 Jun 01.
Artigo em Inglês | MEDLINE | ID: mdl-34206129

RESUMO

Endometriosis is a common disease. Its pathogenesis still remains uncertain, but it is clear that cell proliferation, apoptosis and chronic inflammation play an important role in its development. This paper aimed to investigate the anti-proliferative and anti-inflammatory effects of a combined therapy with fotemustine and dexamethasone. Endometriosis was induced by intraperitoneal injections of uterine fragments from donor animals to recipient animals. Next, the pathology was allowed to develop for 7 days. On the seventh day, fotemustine was administered once and dexamethasone was administered daily for the next 7 days. On Day 14, the animals were sacrificed, and peritoneal fluids and lesions were explanted. In order to evaluate the gastrointestinal side effects of the drugs, stomachs were harvested as well. The combined therapy of fotemustine and dexamethasone reduced the proinflammatory mediator levels in the peritoneal fluid and reduced the lesions' area and diameter. In particular, fotemustine and dexamethasone administration reduced the heterogeneous development of endometrial stroma and glands (histological analysis of lesions) and hyperproliferation of endometriotic cells (immunohistochemical analysis of Ki67 and Western blot analysis of PCNA) through the mitogen-activated protein kinase (MAPK) signaling pathway. Combined fotemustine and dexamethasone therapy showed anti-inflammatory effects by inducing the synthesis of anti-inflammatory mediators at the transcriptional and post-transcriptional levels (Western blot analysis of NFκB, COX-2 and PGE2 expression). Fotemustine and dexamethasone administration had anti-apoptotic activity, restoring the impaired mechanism (TUNEL assay and Western blot analysis of Bax and Bcl-2). Moreover, no gastric disfunction was detected (histological analysis of stomachs). Thus, our data showed that the combined therapy of fotemustine and dexamethasone reduced endometriosis-induced inflammation, hyperproliferation and apoptosis resistance.


Assuntos
Dexametasona/farmacologia , Endometriose/tratamento farmacológico , Inflamação/tratamento farmacológico , Compostos de Nitrosoureia/farmacologia , Compostos Organofosforados/farmacologia , Animais , Apoptose/efeitos dos fármacos , Líquido Ascítico/metabolismo , Proliferação de Células/efeitos dos fármacos , Endometriose/complicações , Endometriose/genética , Endometriose/patologia , Endométrio/efeitos dos fármacos , Endométrio/patologia , Feminino , Humanos , Inflamação/complicações , Inflamação/genética , Inflamação/patologia , NF-kappa B/genética , Antígeno Nuclear de Célula em Proliferação/genética , Ratos , Transdução de Sinais/efeitos dos fármacos , Fator de Transcrição RelA/genética
3.
PLoS One ; 16(7): e0254167, 2021.
Artigo em Inglês | MEDLINE | ID: mdl-34214123

RESUMO

Dexamethasone provides benefits in patients with coronavirus disease 2019 (COVID-19), although data regarding immunological profiles and viral clearance are limited. This study aimed to evaluate for differences in biomarkers among patients with severe COVID-19 who did and did not receive dexamethasone. We measured plasma biomarkers of lung epithelial/endothelial injury and inflammation in 31 patients with severe COVID-19 and in 13 controls. Changes in biomarkers and clinical parameters were compared during the 7-day period among COVID-19 patients, and also according to dexamethasone use. Thirty-two patients with severe COVID-19 who received mechanical ventilation (n = 6), high-flow nasal cannula (n = 11), and supplemental oxygen (n = 15) were analyzed. Relative to controls, patients with severe COVID-19 had significantly higher concentrations of biomarkers related to glycocalyx shedding (endocan and syndecan-1), endothelial injury (von Willebrand factor), and inflammation (soluble receptor for advanced glycation end-products [sRAGE] and interleukin-6). The 7-day decreases in biomarkers of endothelial injury (angiopoietin-2 [Ang-2] and intercellular adhesion molecule-1 [ICAM-1]) and sRAGE, but not in the biomarker of lung epithelial injury (surfactant protein D), were correlated with decreases in C-reactive protein and radiologic score at day 7. Twenty patients (63%) received dexamethasone, and the dexamethasone and non-dexamethasone groups differed in terms of disease severity. However, dexamethasone was associated marginally with increased SpO2/FiO2 and significantly with decreases in C-reactive protein and radiologic score after adjusting for baseline imbalances. Furthermore, the dexamethasone group exhibited a significant decrease in the concentrations of Ang-2, ICAM-1, soluble form of the Tie2 receptor (a biomarker of glycocalyx shedding), and sRAGE. Both groups exhibited a clinically insignificant increase in the cycle threshold value. Severe COVID-19 may be characterized by more severe endothelial injury and inflammation, and less severe lung epithelial injury. There is a possibility that dexamethasone improved severe COVID-19 and related endothelial injury without delaying viral clearance.


Assuntos
Anti-Inflamatórios/uso terapêutico , COVID-19/tratamento farmacológico , Dexametasona/uso terapêutico , Endotélio Vascular/efeitos dos fármacos , Inflamação/prevenção & controle , SARS-CoV-2 , Viremia/tratamento farmacológico , Adulto , Idoso , Anti-Inflamatórios/farmacologia , Biomarcadores , COVID-19/sangue , COVID-19/diagnóstico por imagem , Dexametasona/farmacologia , Endotélio Vascular/patologia , Feminino , Humanos , Inflamação/sangue , Inflamação/etiologia , Lesão Pulmonar/sangue , Lesão Pulmonar/diagnóstico por imagem , Lesão Pulmonar/etiologia , Masculino , Oxigênio/sangue , Projetos Piloto , Carga Viral , Viremia/sangue
4.
Int J Mol Sci ; 22(13)2021 Jun 23.
Artigo em Inglês | MEDLINE | ID: mdl-34201797

RESUMO

The coronavirus disease 2019 (COVID-19) caused by infection of the severe respiratory syndrome coronavirus-2 (SARS-CoV-2) significantly impacted human society. Recently, the synthetic pure glucocorticoid dexamethasone was identified as an effective compound for treatment of severe COVID-19. However, glucocorticoids are generally harmful for infectious diseases, such as bacterial sepsis and severe influenza pneumonia, which can develop respiratory failure and systemic inflammation similar to COVID-19. This apparent inconsistency suggests the presence of pathologic mechanism(s) unique to COVID-19 that renders this steroid effective. We review plausible mechanisms and advance the hypothesis that SARS-CoV-2 infection is accompanied by infected cell-specific glucocorticoid insensitivity as reported for some other viruses. This alteration in local glucocorticoid actions interferes with undesired glucocorticoid to facilitate viral replication but does not affect desired anti-inflammatory properties in non-infected organs/tissues. We postulate that the virus coincidentally causes glucocorticoid insensitivity in the process of modulating host cell activities for promoting its replication in infected cells. We explore this tenet focusing on SARS-CoV-2-encoding proteins and potential molecular mechanisms supporting this hypothetical glucocorticoid insensitivity unique to COVID-19 but not characteristic of other life-threatening viral diseases, probably due to a difference in specific virally-encoded molecules and host cell activities modulated by them.


Assuntos
Antivirais/farmacologia , COVID-19/tratamento farmacológico , Dexametasona/farmacologia , Sistema Hipotálamo-Hipofisário/fisiologia , Inflamação/tratamento farmacológico , Interações entre Hospedeiro e Microrganismos , Humanos , Imunidade Inata , SARS-CoV-2/efeitos dos fármacos , SARS-CoV-2/fisiologia , Índice de Gravidade de Doença , Replicação Viral/efeitos dos fármacos
5.
Int J Mol Sci ; 22(10)2021 May 20.
Artigo em Inglês | MEDLINE | ID: mdl-34065385

RESUMO

In retinitis pigmentosa (RP), one of many possible genetic mutations causes rod degeneration, followed by cone secondary death leading to blindness. Accumulating evidence indicates that rod death triggers multiple, non-cell-autonomous processes, which include oxidative stress and inflammation/immune responses, all contributing to cone demise. Inflammation relies on local microglia and recruitment of immune cells, reaching the retina through breakdowns of the inner blood retinal barrier (iBRB). Leakage in the inner retina vasculature suggests similarly altered outer BRB, formed by junctions between retinal pigment epithelium (RPE) cells, which are crucial for retinal homeostasis, immune response, and privilege. We investigated the RPE structural integrity in three models of RP (rd9, rd10, and Tvrm4 mice) by immunostaining for zonula occludens-1 (ZO-1), an essential regulatory component of tight junctions. Quantitative image analysis demonstrated discontinuities in ZO-1 profiles in all mutants, despite different degrees of photoreceptor loss. ZO-1 interruption zones corresponded to leakage of in vivo administered, fluorescent dextran through the choroid-RPE interface, demonstrating barrier dysfunction. Dexamethasone, administered to rd10 mice for rescuing cones, also rescued RPE structure. Thus, previously undetected, stereotyped abnormalities occur in the RPE of RP mice; pharmacological targeting of inflammation supports a feedback loop leading to simultaneous protection of cones and the RPE.


Assuntos
Retina/fisiopatologia , Epitélio Pigmentado da Retina/fisiopatologia , Retinite Pigmentosa/fisiopatologia , Animais , Dexametasona/farmacologia , Modelos Animais de Doenças , Estudos de Avaliação como Assunto , Inflamação/metabolismo , Inflamação/fisiopatologia , Camundongos , Camundongos Endogâmicos C57BL , Retina/efeitos dos fármacos , Retina/metabolismo , Células Fotorreceptoras Retinianas Cones/efeitos dos fármacos , Células Fotorreceptoras Retinianas Cones/metabolismo , Epitélio Pigmentado da Retina/efeitos dos fármacos , Epitélio Pigmentado da Retina/metabolismo , Células Fotorreceptoras Retinianas Bastonetes/efeitos dos fármacos , Células Fotorreceptoras Retinianas Bastonetes/metabolismo , Vasos Retinianos/efeitos dos fármacos , Vasos Retinianos/metabolismo , Retinite Pigmentosa/metabolismo , Rodopsina/metabolismo , Junções Íntimas/efeitos dos fármacos , Junções Íntimas/metabolismo , Proteína da Zônula de Oclusão-1/metabolismo
6.
Blood Adv ; 5(12): 2593-2607, 2021 06 22.
Artigo em Inglês | MEDLINE | ID: mdl-34152396

RESUMO

Novel combination therapies have markedly improved the lifespan of patients with multiple myeloma (MM), but drug resistance and disease relapse remain major clinical problems. Dexamethasone and other glucocorticoids are a cornerstone of conventional and new combination therapies for MM, although their use is accompanied by serious side effects. We aimed to uncover drug combinations that act in synergy and, as such, allow reduced dosing while remaining effective. Dexamethasone and the myeloid cell leukemia 1 (MCL-1) inhibitor S63845 (MCL-1i) proved the most potent combination in our lethality screen and induced apoptosis of human myeloma cell lines (HMCLs) that was 50% higher compared with an additive drug effect. Kinome analysis of dexamethasone-treated HMCLs revealed a reduction in serine/threonine peptide phosphorylation, which was predicted to result from reduced Akt activity. Biochemical techniques showed no dexamethasone-induced effects on FOXO protein or GSK3 but did show a 50% reduction in P70S6K phosphorylation, downstream of the Akt-mTORC1 axis. Replacing dexamethasone by the P70S6K1 isoform-specific inhibitor PF-4708671 (S6K1i) revealed similar and statistically significant synergistic apoptosis of HMCLs in combination with MCL-1i. Interestingly, apoptosis induced by the P70S6K1i and MCL-1i combination was more-than-additive in all 9 primary MM samples tested; this effect was observed for 6 of 9 samples with the dexamethasone and MCL-1i combination. Toxicity on stem and progenitor cell subsets remained minimal. Combined, our results show a strong rationale for combination treatments using the P70S6K inhibitor in MM. Direct and specific inhibition of P70S6K may also provide a solution for patients ineligible or insensitive to dexamethasone or other glucocorticoids.


Assuntos
Mieloma Múltiplo , Linhagem Celular Tumoral , Dexametasona/farmacologia , Quinase 3 da Glicogênio Sintase , Humanos , Mieloma Múltiplo/tratamento farmacológico , Proteína de Sequência 1 de Leucemia de Células Mieloides , Proteínas Quinases S6 Ribossômicas 70-kDa
7.
Per Med ; 18(4): 389-398, 2021 07.
Artigo em Inglês | MEDLINE | ID: mdl-34086487

RESUMO

Immunomodulatory and analgesic effects of dexamethasone are clinically well established, and this synthetic corticosteroid acts as an agonist of glucocorticoid receptors. Early results of the RECOVERY Trial from the United Kingdom and others suggest certain benefits of dexamethasone against COVID-19 chronic patients. The efforts have been acknowledged by World Health Organization with an interim guideline to use in patients with a severe and critical illness. The inherent genetic variations in genes such as CYP3A5, NR3C1, NR3C2, etc., involved in the pharmacokinetic and pharmacodynamic processes may influence dexamethasone's effects as an anti-inflammatory drug. Besides, the drug may influence transcriptome or metabolic changes in the individuals. In the present review, we summarize the reported genetic variations that impact dexamethasone response and discuss dexamethasone-induced changes in transcriptome and metabolome that may influence potential treatment outcome against COVID-19.


Assuntos
COVID-19/tratamento farmacológico , COVID-19/genética , Dexametasona/farmacologia , Reposicionamento de Medicamentos , Glucocorticoides/uso terapêutico , Farmacogenética , SARS-CoV-2 , Animais , Feminino , Frequência do Gene , Variação Genética , Humanos , Masculino , Metaboloma , Modelos Animais , Preparações Farmacêuticas , Guias de Prática Clínica como Assunto , Transcriptoma
8.
Int J Mol Sci ; 22(11)2021 May 31.
Artigo em Inglês | MEDLINE | ID: mdl-34072647

RESUMO

Steroid-induced glaucoma is a severe pathological condition, sustained by a rapidly progressive increase in intraocular pressure (IOP), which is diagnosed in a subset of subjects who adhere to a glucocorticoid (GC)-based therapy. Molecular and clinical studies suggest that either natural or synthetic GCs induce a severe metabolic dysregulation of Trabecular Meshwork Cells (TMCs), an endothelial-derived histotype with phagocytic and secretive functions which lay at the iridocorneal angle in the anterior segment of the eye. Since TMCs physiologically regulate the composition and architecture of trabecular meshwork (TM), which is the main outflow pathway of aqueous humor, a fluid which shapes the eye globe and nourishes the lining cell types, GCs are supposed to trigger a pathological remodeling of the TM, inducing an IOP increase and retina mechanical compression. The metabolic dysregulation of TMCs induced by GCs exposure has never been characterized at the molecular detail. Herein, we report that, upon dexamethasone exposure, a TMCs strain develops a marked inhibition of the autophagosome biogenesis pathway through an enhanced turnover of two members of the Ulk-1 complex, the main platform for autophagy induction, through the Ubiquitin Proteasome System (UPS).


Assuntos
Proteína Homóloga à Proteína-1 Relacionada à Autofagia/metabolismo , Autofagia/efeitos dos fármacos , Dexametasona/farmacologia , Peptídeos e Proteínas de Sinalização Intracelular/metabolismo , Complexos Multiproteicos/metabolismo , Malha Trabecular/efeitos dos fármacos , Malha Trabecular/metabolismo , Apoptose/efeitos dos fármacos , Autofagia/genética , Proteína Homóloga à Proteína-1 Relacionada à Autofagia/genética , Proliferação de Células/efeitos dos fármacos , Dexametasona/efeitos adversos , Suscetibilidade a Doenças , Glaucoma/etiologia , Glaucoma/metabolismo , Glaucoma/fisiopatologia , Humanos , Peptídeos e Proteínas de Sinalização Intracelular/genética , Complexo de Endopeptidases do Proteassoma/metabolismo
9.
Int J Mol Sci ; 22(11)2021 May 27.
Artigo em Inglês | MEDLINE | ID: mdl-34072036

RESUMO

The hypothalamic-pituitary-adrenal axis is stimulated in response to stress. When activated, it is suppressed by the negative feedback effect of glucocorticoids. Glucocorticoids directly inhibit proopiomelanocortin (Pomc) gene expression in the pituitary. Glucocorticoid signaling is mediated via glucocorticoid receptors, 11ß-hydroxysteroid dehydrogenases, and the FK506-binding immunophilins, Fkbp4 and Fkbp5. Fkbp4 and Fkbp5 differentially regulate dynein interaction and nuclear translocation of the glucocorticoid receptor, resulting in modulation of the glucocorticoid action. Here, we explored the regulation of Fkbp4 and Fkbp5 genes and their proteins with dexamethasone, a major synthetic glucocorticoid drug, in murine AtT-20 corticotroph cells. To elucidate further roles of Fkbp4 and Fkbp5, we examined their effects on Pomc mRNA levels in corticotroph cells. Dexamethasone decreased Pomc mRNA levels as well as Fkpb4 mRNA levels in mouse corticotroph cells. Dexamethasone tended to decrease Fkbp4 protein levels, while it increased Fkpb5 mRNA and its protein levels. The dexamethasone-induced decreases in Pomc mRNA levels were partially canceled by Fkbp4 knockdown. Alternatively, Pomc mRNA levels were further decreased by Fkbp5 knockdown. Thus, Fkbp4 contributes to the negative feedback of glucocorticoids, and Fkbp5 reduces the efficiency of the glucocorticoid effect on Pomc gene expression in pituitary corticotroph cells.


Assuntos
Corticotrofos/metabolismo , Regulação da Expressão Gênica , Pró-Opiomelanocortina/genética , Proteínas de Ligação a Tacrolimo/metabolismo , Animais , Biomarcadores , Corticotrofos/citologia , Dexametasona/farmacologia , Regulação da Expressão Gênica/efeitos dos fármacos , Técnicas de Silenciamento de Genes , Glucocorticoides/metabolismo , Camundongos , Modelos Biológicos , Ligação Proteica , RNA Mensageiro/genética , Receptores de Glucocorticoides/metabolismo , Transdução de Sinais/efeitos dos fármacos , Proteínas de Ligação a Tacrolimo/genética
10.
Int J Mol Sci ; 22(10)2021 May 14.
Artigo em Inglês | MEDLINE | ID: mdl-34069142

RESUMO

Bone healing is a complex, well-organized process. Multiple factors regulate this process, including growth factors, hormones, cytokines, mechanical stimulation, and aging. One of the most important signaling pathways that affect bone healing is the Notch signaling pathway. It has a significant role in controlling the differentiation of bone mesenchymal stem cells and forming new bone. Interventions to enhance the healing of critical-sized bone defects are of great importance, and stem cell transplantations are eminent candidates for treating such defects. Understanding how Notch signaling impacts pluripotent stem cell differentiation can significantly enhance osteogenesis and improve the overall healing process upon transplantation. In Rancourt's lab, mouse embryonic stem cells (ESC) have been successfully differentiated to the osteogenic cell lineage. This study investigates the role of Notch signaling inhibition in the osteogenic differentiation of mouse embryonic and induced pluripotent stem cells (iPS). Our data showed that Notch inhibition greatly enhanced the differentiation of both mouse embryonic and induced pluripotent stem cells.


Assuntos
Secretases da Proteína Precursora do Amiloide/antagonistas & inibidores , Diferenciação Celular/efeitos dos fármacos , Osteogênese/genética , Células-Tronco Pluripotentes/citologia , Células-Tronco Pluripotentes/efeitos dos fármacos , Animais , Osso e Ossos/metabolismo , Proteínas de Ciclo Celular/genética , Diferenciação Celular/fisiologia , Subunidade alfa 1 de Fator de Ligação ao Core/genética , Subunidade alfa 1 de Fator de Ligação ao Core/metabolismo , Dexametasona/farmacologia , Diaminas/farmacologia , Regulação da Expressão Gênica/efeitos dos fármacos , Células-Tronco Pluripotentes Induzidas/citologia , Células-Tronco Pluripotentes Induzidas/efeitos dos fármacos , Células-Tronco Pluripotentes Induzidas/fisiologia , Mesoderma/citologia , Camundongos , Células-Tronco Embrionárias Murinas/citologia , Células-Tronco Embrionárias Murinas/efeitos dos fármacos , Osteoblastos/citologia , Osteoblastos/efeitos dos fármacos , Osteoblastos/fisiologia , Osteogênese/efeitos dos fármacos , Células-Tronco Pluripotentes/metabolismo , Receptores Notch/metabolismo , Tiazóis/farmacologia , Fatores de Transcrição HES-1/genética , Vitamina D/farmacologia
11.
Zhongguo Dang Dai Er Ke Za Zhi ; 23(6): 639-644, 2021 Jun.
Artigo em Chinês | MEDLINE | ID: mdl-34130788

RESUMO

OBJECTIVE: To study the effect of dexamethasone (DEX) on the expression of Dynein heavy chain (DHC) and Dynactin in the cytoplasm of fetal rat cerebral cortical neurons cultured in vitro. METHODS: Primary cerebral cortical neurons of fetal rats were cultured in vitro and were used to establish a cellular model of DEX intervention. According to the final concentration of DEX, the neurons were divided into three groups:control (without DEX), 0.1 µmol/L DEX, and 1.0 µmol/L DEX. On days 1, 3, and 7 after intervention, the quantitative PCR was used to observe the effect of DEX on the mRNA expression of DHC and Dynactin. The Western blot was used to observe the effect of DEX on the protein expression of DHC and Dynactin. RESULTS: There was no significant difference in the mRNA expression levels of DHC and Dynactin among the three groups at all time points (P > 0.05). On day 7 after DEX intervention, the protein expression of DHC in the 1.0 µmol/L DEX group gradually increased and reached the peak over time, which was significantly higher than that in the control and 0.1 µmol/L DEX groups (P < 0.05). The control and 0.1 µmol/L DEX groups had a significant increase in the protein expression of Dynactin from day 1 to days 3 and 7 after DEX intervention (P < 0.05). The control group had a significant increase in the protein expression of Dynactin from day 3 to day 7 after intervention (P < 0.05), while the 0.1 µmol/L DEX group had a significant reduction in the protein expression of Dynactin from day 3 to day 7 after intervention (P < 0.05). On days 3 and 7 after DEX intervention, the 0.1 µmol/L DEX and 1.0 µmol/L DEX groups had a significantly lower protein expression level of Dynactin in the cerebral cortical neurons than the control group (P < 0.05). On day 7 after DEX intervention, the 1.0 µmol/L DEX group had a significantly lower protein expression level of Dynactin than the 0.1 µmol/L DEX group (P < 0.05). CONCLUSIONS: DEX affects the protein expression of DHC and Dynactin in the fetal rat cerebral cortical neurons cultured in vitro, possibly in a concentration- and time-dependent manner.


Assuntos
Dineínas , Neurônios , Animais , Citoplasma , Dexametasona/farmacologia , Complexo Dinactina/genética , Ratos
12.
Life Sci ; 278: 119564, 2021 Aug 01.
Artigo em Inglês | MEDLINE | ID: mdl-33961857

RESUMO

AIMS: Elevated intraocular pressure is primarily induced by the increased resistance of conventional outflow of aqueous humor. Dysfunction of the juxtacanalicular region of trabecular meshwork (TM) and Schlemm's canal (SC) endothelium, as the main conventional outflow tissue, have been implicated as the major reasons for the increased resistance. Integrins are widespread in these tissues, especially alpha8 integrin (ITGA8). We aim to investigate the properties of cells expressing ITGA8 in the conventional outflow tissue. MAIN METHODS: Fluorescence in situ hybridization (FISH) and immunofluorescence (IF) were performed to detect the mRNA and protein levels of ITGA8 in human conventional outflow tissue. ITGA8-positive cells were isolated from the cultured human TM cells through a magnetic bead-based approach. Flow Cytometry was used to determine the purification efficiency. The expressions of TM and SC biomarkers and dexamethasone-induced myocilin secretion capacity of ITGA8-positive cells was assessed by Real-time PCR, IF and Western blot. A gel contraction assay was performed to evaluate contractility of ITGA8-positive cells after endothelin 1 treatment. KEY FINDINGS: ITGA8 was found with robust expression near the inner wall of SC endothelium. After purification, the proportion of ITGA8-positive cells were increased by about 10%. ITGA8-positive cells were identified with the properties as SC endothelial cells, such as more robust expressions of SC biomarkers, less dexamethasone-inducible myocilin expression, and stronger contractility. SIGNIFICANCE: This study demonstrated that cells expressing ITGA8 in SC region possess more properties as SC endothelial cells. Our data implicate a crucial role of ITGA8 in aqueous humor (AH) outflow resistance regulation.


Assuntos
Humor Aquoso/metabolismo , Células Endoteliais/metabolismo , Glaucoma/metabolismo , Cadeias alfa de Integrinas/metabolismo , Pressão Intraocular , Malha Trabecular/metabolismo , Biomarcadores/metabolismo , Sobrevivência Celular , Proteínas do Citoesqueleto/metabolismo , Dexametasona/farmacologia , Endotelina-1/metabolismo , Endotélio/metabolismo , Proteínas do Olho/metabolismo , Glaucoma/fisiopatologia , Glucocorticoides/metabolismo , Glicoproteínas/metabolismo , Humanos , Hibridização in Situ Fluorescente , Integrinas/metabolismo
13.
Stem Cell Reports ; 16(5): 1165-1181, 2021 05 11.
Artigo em Inglês | MEDLINE | ID: covidwho-1225410

RESUMO

SARS-CoV-2 infection is associated with lower blood oxygen levels, even in patients without hypoxia requiring hospitalization. This discordance illustrates the need for a more unifying explanation as to whether SARS-CoV-2 directly or indirectly affects erythropoiesis. Here, we show significantly enriched CD71+ erythroid precursors/progenitors in the blood circulation of COVID-19 patients. We found that these cells have distinctive immunosuppressive properties. In agreement, we observed a strong negative correlation between the frequency of these cells with T and B cell proportions in COVID-19 patients. The expansion of these CD71+ erythroid precursors/progenitors was negatively correlated with the hemoglobin levels. A subpopulation of abundant erythroid cells, CD45+ CD71+ cells, co-express ACE2, TMPRSS2, CD147, and CD26, and these can be infected with SARS-CoV-2. In turn, pre-treatment of erythroid cells with dexamethasone significantly diminished ACE2/TMPRSS2 expression and subsequently reduced their infectivity with SARS-CoV-2. This provides a novel insight into the impact of SARS-CoV-2 on erythropoiesis and hypoxia seen in COVID-19 patients.


Assuntos
Imunidade Adaptativa/imunologia , COVID-19/patologia , Células Precursoras Eritroides/virologia , Eritropoese/fisiologia , Hemoglobinas/análise , Oxigênio/sangue , Adolescente , Adulto , Idoso , Idoso de 80 Anos ou mais , Enzima de Conversão de Angiotensina 2/metabolismo , Animais , Linfócitos B/citologia , Linfócitos B/imunologia , COVID-19/imunologia , Dexametasona/farmacologia , Células Precursoras Eritroides/imunologia , Feminino , Humanos , Contagem de Linfócitos , Masculino , Camundongos , Camundongos Endogâmicos BALB C , Pessoa de Meia-Idade , SARS-CoV-2/imunologia , Serina Endopeptidases/metabolismo , Linfócitos T/citologia , Linfócitos T/imunologia , Adulto Jovem
14.
Stem Cell Res Ther ; 12(1): 291, 2021 05 17.
Artigo em Inglês | MEDLINE | ID: mdl-34001245

RESUMO

BACKGROUND: Glucocorticoid-induced osteoporosis (GIOP) is the most common secondary osteoporosis. Patients with GIOP are susceptible to fractures and the subsequent delayed bone union or nonunion. Thus, effective drugs and targets need to be explored. In this regard, the present study aims to reveal the possible mechanism of the anti-GIOP effect of all-trans retinoic acid (ATRA). METHODS: Bone morphogenetic protein 9 (BMP9)-transfected mesenchymal stem cells (MSCs) were used as an in vitro osteogenic model to deduce the relationship between ATRA and dexamethasone (DEX). The osteogenic markers runt-related transcription factor 2 (RUNX2), alkaline phosphatase (ALP), and osteopontin were detected using real-time quantitative polymerase chain reaction, Western blot, and immunofluorescent staining assay. ALP activities and matrix mineralization were evaluated using ALP staining and Alizarin Red S staining assay, respectively. The novel genes associated with ATRA and DEX were detected using RNA sequencing (RNA-seq). The binding of the protein-DNA complex was validated using chromatin immunoprecipitation (ChIP) assay. Rat GIOP models were constructed using intraperitoneal injection of dexamethasone at a dose of 1 mg/kg, while ATRA intragastric administration was applied to prevent and treat GIOP. These effects were evaluated based on the serum detection of the osteogenic markers osteocalcin and tartrate-resistant acid phosphatase 5b, histological staining, and micro-computed tomography analysis. RESULTS: ATRA enhanced BMP9-induced ALP, RUNX2 expressions, ALP activities, and matrix mineralization in mouse embryonic fibroblasts as well as C3H10T1/2 and C2C12 cells, while a high concentration of DEX attenuated these markers. When DEX was combined with ATRA, the latter reversed DEX-inhibited ALP activities and osteogenic markers. In vivo analysis showed that ATRA reversed DEX-inhibited bone volume, bone trabecular number, and thickness. During the reversal process of ATRA, the expression of retinoic acid receptor beta (RARß) was elevated. RARß inhibitor Le135 partly blocked the reversal effect of ATRA. Meanwhile, RNA-seq demonstrated that serine protease inhibitor, clade A, member 3N (Serpina3n) was remarkably upregulated by DEX but downregulated when combined with ATRA. Overexpression of Serpina3n attenuated ATRA-promoted osteogenic differentiation, whereas knockdown of Serpina3n blocked DEX-inhibited osteogenic differentiation. Furthermore, ChIP assay revealed that RARß can regulate the expression of Serpina3n. CONCLUSION: ATRA can reverse DEX-inhibited osteogenic differentiation both in vitro and in vivo, which may be closely related to the downregulation of DEX-promoted Serpina3n. Hence, ATRA may be viewed as a novel therapeutic agent, and Serpina3n may act as a new target for GIOP.


Assuntos
Células-Tronco Mesenquimais , Serpinas , Proteínas de Fase Aguda , Animais , Diferenciação Celular , Células Cultivadas , Dexametasona/farmacologia , Fibroblastos , Humanos , Camundongos , Osteogênese , Ratos , Tretinoína/farmacologia , Microtomografia por Raio-X
15.
Int J Mol Sci ; 22(9)2021 Apr 29.
Artigo em Inglês | MEDLINE | ID: mdl-33947115

RESUMO

Cortisol, a stress hormone, plays key roles in mediating stress and anti-inflammatory responses. As abnormal cortisol levels can induce various adverse effects, screening cortisol and cortisol analogues is important for monitoring stress levels and for identifying drug candidates. A novel cell-based sensing system was adopted for rapid screening of cortisol and its functional analogues under complex cellular regulation. We used glucocorticoid receptor (GR) fused to a split intein which reconstituted with the counterpart to trigger conditional protein splicing (CPS) in the presence of targets. CPS generates functional signal peptides which promptly translocate the fluorescent cargo. The sensor cells exhibited exceptional performance in discriminating between the functional and structural analogues of cortisol with improved sensitivity. Essential oil extracts with stress relief activity were screened using the sensor cells to identify GR effectors. The sensor cells responded to peppermint oil, and L-limonene and L-menthol were identified as potential GR effectors from the major components of peppermint oil. Further analysis indicated L-limonene as a selective GR agonist (SEGRA) which is a potential anti-inflammatory agent as it attenuates proinflammatory responses without causing notable adverse effects of GR agonists.


Assuntos
Técnicas Biossensoriais , Avaliação Pré-Clínica de Medicamentos/métodos , Polarização de Fluorescência/métodos , Hidrocortisona/análise , Óleos Voláteis/farmacologia , Receptores de Glucocorticoides/agonistas , Atrofia , Acetato de Ciproterona/farmacologia , Dexametasona/farmacologia , Estradiol/farmacologia , Fluorometria , Células HeLa , Humanos , Inteínas , Limoneno/farmacologia , Proteínas Luminescentes/análise , Mentol/farmacologia , Mifepristona/farmacologia , Estrutura Molecular , Músculo Esquelético/patologia , Mioblastos/efeitos dos fármacos , Óleos Vegetais/farmacologia , Processamento de Proteína , Transporte Proteico , Proteínas Recombinantes de Fusão/genética , Proteínas Recombinantes de Fusão/metabolismo
16.
Stem Cell Reports ; 16(5): 1165-1181, 2021 05 11.
Artigo em Inglês | MEDLINE | ID: mdl-33979601

RESUMO

SARS-CoV-2 infection is associated with lower blood oxygen levels, even in patients without hypoxia requiring hospitalization. This discordance illustrates the need for a more unifying explanation as to whether SARS-CoV-2 directly or indirectly affects erythropoiesis. Here, we show significantly enriched CD71+ erythroid precursors/progenitors in the blood circulation of COVID-19 patients. We found that these cells have distinctive immunosuppressive properties. In agreement, we observed a strong negative correlation between the frequency of these cells with T and B cell proportions in COVID-19 patients. The expansion of these CD71+ erythroid precursors/progenitors was negatively correlated with the hemoglobin levels. A subpopulation of abundant erythroid cells, CD45+ CD71+ cells, co-express ACE2, TMPRSS2, CD147, and CD26, and these can be infected with SARS-CoV-2. In turn, pre-treatment of erythroid cells with dexamethasone significantly diminished ACE2/TMPRSS2 expression and subsequently reduced their infectivity with SARS-CoV-2. This provides a novel insight into the impact of SARS-CoV-2 on erythropoiesis and hypoxia seen in COVID-19 patients.


Assuntos
Imunidade Adaptativa/imunologia , COVID-19/patologia , Células Precursoras Eritroides/virologia , Eritropoese/fisiologia , Hemoglobinas/análise , Oxigênio/sangue , Adolescente , Adulto , Idoso , Idoso de 80 Anos ou mais , Enzima de Conversão de Angiotensina 2/metabolismo , Animais , Linfócitos B/citologia , Linfócitos B/imunologia , COVID-19/imunologia , Dexametasona/farmacologia , Células Precursoras Eritroides/imunologia , Feminino , Humanos , Contagem de Linfócitos , Masculino , Camundongos , Camundongos Endogâmicos BALB C , Pessoa de Meia-Idade , SARS-CoV-2/imunologia , Serina Endopeptidases/metabolismo , Linfócitos T/citologia , Linfócitos T/imunologia , Adulto Jovem
17.
Int J Mol Sci ; 22(9)2021 Apr 30.
Artigo em Inglês | MEDLINE | ID: mdl-33946412

RESUMO

Despite the huge body of research on osteogenic differentiation and bone tissue engineering, the translation potential of in vitro results still does not match the effort employed. One reason might be that the protocols used for in vitro research have inherent pitfalls. The synthetic glucocorticoid dexamethasone is commonly used in protocols for trilineage differentiation of human bone marrow mesenchymal stromal cells (hBMSCs). However, in the case of osteogenic commitment, dexamethasone has the main pitfall of inhibiting terminal osteoblast differentiation, and its pro-adipogenic effect is well known. In this work, we aimed to clarify the role of dexamethasone in the osteogenesis of hBMSCs, with a particular focus on off-target differentiation. The results showed that dexamethasone does induce osteogenic differentiation by inhibiting SOX9 expression, but not directly through RUNX2 upregulation as it is commonly thought. Rather, PPARG is concomitantly and strongly upregulated, leading to the formation of adipocyte-like cells within osteogenic cultures. Limiting the exposure to dexamethasone to the first week of differentiation did not affect the mineralization potential. Gene expression levels of RUNX2, SOX9, and PPARG were simulated using approximate Bayesian computation based on a simplified theoretical model, which was able to reproduce the observed experimental trends but with a different range of responses, indicating that other factors should be integrated to fully understand how dexamethasone influences cell fate. In summary, this work provides evidence that current in vitro differentiation protocols based on dexamethasone do not represent a good model, and further research is warranted in this field.


Assuntos
Dexametasona/farmacologia , Glucocorticoides/farmacologia , Células-Tronco Mesenquimais/efeitos dos fármacos , Osteogênese/efeitos dos fármacos , PPAR gama/metabolismo , Fatores de Transcrição SOX9/metabolismo , Adulto , Diferenciação Celular/efeitos dos fármacos , Células Cultivadas , Subunidade alfa 1 de Fator de Ligação ao Core/genética , Subunidade alfa 1 de Fator de Ligação ao Core/metabolismo , Feminino , Regulação da Expressão Gênica/efeitos dos fármacos , Humanos , Masculino , Células-Tronco Mesenquimais/citologia , Células-Tronco Mesenquimais/metabolismo , PPAR gama/genética , Fatores de Transcrição SOX9/genética
18.
Int J Mol Sci ; 22(7)2021 Mar 31.
Artigo em Inglês | MEDLINE | ID: mdl-33807481

RESUMO

Glucocorticoids (GCs) act via the GC receptor (GR), a receptor ubiquitously expressed in the body where it drives a broad spectrum of responses within distinct cell types and tissues, which vary in strength and specificity. The variability of GR-mediated cell responses is further extended by the existence of GR isoforms, such as GRα and GRß, generated through alternative splicing mechanisms. While GRα is the classic receptor responsible for GC actions, GRß has been implicated in the impairment of GRα-mediated activities. Interestingly, in contrast to the popular belief that GRß actions are restricted to its dominant-negative effects on GRα-mediated responses, GRß has been shown to have intrinsic activities and "directly" regulates a plethora of genes related to inflammatory process, cell communication, migration, and malignancy, each in a GRα-independent manner. Furthermore, GRß has been associated with increased cell migration, growth, and reduced sensitivity to GC-induced apoptosis. We will summarize the current knowledge of GRß-mediated responses, with a focus on the GRα-independent/intrinsic effects of GRß and the associated non-canonical signaling pathways. Where appropriate, potential links to airway inflammatory diseases will be highlighted.


Assuntos
Receptores de Glucocorticoides/metabolismo , Receptores de Glucocorticoides/fisiologia , Processamento Alternativo/efeitos dos fármacos , Dexametasona/farmacologia , Glucocorticoides/farmacologia , Humanos , Isoformas de Proteínas
19.
Mol Med Rep ; 23(5)2021 05.
Artigo em Inglês | MEDLINE | ID: mdl-33649814

RESUMO

Muscle atrophy, a side effect from administration of the anti­inflammatory medication dexamethasone (DEX), is preventable by concomitant administration of the major monomeric constituent of Panax ginseng C.A. Meyer, 20(S)­ginsenoside Rg3 (S­Rg3). Putative S­Rg3­associated prevention of DEX­induced muscle atrophy may involve S­Rg3 mitigation of DEX­induced mitochondrial dysfunction. In the present study, MTT assays revealed enhanced cell viability following S­Rg3 treatment of DEX­injured C2C12 myotubes. Subsequent PCR and western blotting results demonstrated S­Rg3­induced reduction of expression of muscle atrophy F­box protein (atrogin­1) and muscle RING­finger protein­1, proteins previously linked to muscle atrophy. Additionally, S­Rg3 treatment of DEX­injured myotubes led to aggregation of Rg3 monomers in cells and dose­dependent increases in cellular mitochondrial basal respiratory oxygen consumption rate and intracellular ATP levels compared with their levels in untreated DEX­injured myotubes. In addition, S­Rg3 treatment significantly reversed DEX­induced reductions of expression of key mitochondrial respiratory electron transport chain subunits of protein complexes II, III and V in DEX­injured myotube cells. Furthermore, S­Rg3 alleviation of mitochondrial dysfunction associated with DEX­induced injury of C2C12 myotubes was linked to S­Rg3­associated decreases in both forkhead box O3 (FoxO3) protein expression and phosphorylation of AMP­activated protein kinase (AMPK). Collectively, these results implicate S­Rg3 modulation of signaling within the AMPK­FoxO3 pathway as a putative mechanism underlying S­Rg3 alleviation of DEX­induced muscle atrophy.


Assuntos
Proteínas Quinases Ativadas por AMP/genética , Dexametasona/farmacologia , Proteína Forkhead Box O3/genética , Ginsenosídeos/farmacologia , Mitocôndrias Musculares/efeitos dos fármacos , Fibras Musculares Esqueléticas/efeitos dos fármacos , Transdução de Sinais/efeitos dos fármacos , Proteínas Quinases Ativadas por AMP/metabolismo , Trifosfato de Adenosina/metabolismo , Animais , Linhagem Celular , Sobrevivência Celular/efeitos dos fármacos , Sobrevivência Celular/genética , Complexo de Proteínas da Cadeia de Transporte de Elétrons/genética , Complexo de Proteínas da Cadeia de Transporte de Elétrons/metabolismo , Proteína Forkhead Box O3/metabolismo , Regulação da Expressão Gênica/efeitos dos fármacos , Glucocorticoides/farmacologia , Camundongos , Mitocôndrias Musculares/metabolismo , Modelos Biológicos , Fibras Musculares Esqueléticas/citologia , Fibras Musculares Esqueléticas/metabolismo , Proteínas Musculares/genética , Proteínas Musculares/metabolismo , Atrofia Muscular/genética , Atrofia Muscular/metabolismo , Atrofia Muscular/prevenção & controle , Proteínas Ligases SKP Culina F-Box/genética , Proteínas Ligases SKP Culina F-Box/metabolismo , Transdução de Sinais/genética , Proteínas com Motivo Tripartido/genética , Proteínas com Motivo Tripartido/metabolismo , Ubiquitina-Proteína Ligases/genética , Ubiquitina-Proteína Ligases/metabolismo
20.
Res Vet Sci ; 136: 182-184, 2021 May.
Artigo em Inglês | MEDLINE | ID: mdl-33676156

RESUMO

Anesthetics may worsen the outcome of patients undergoing oncologic surgery via immune suppression. This study examines the impact of propofol, ketamine, and alfaxalone on canine peripheral blood lymphocyte (PBL) cytotoxic function in vitro. PBLs isolated from healthy canine blood were cultured in the presence or absence of alfaxalone, propofol, ketamine, their carrier solutions or dexamethasone as a positive control for 20 h. There was a decrease in cytotoxicity in PBLs exposed to dexamethasone and propofol carrier compared to the control as assessed by means of a chromium-based cytotoxicity assay. The PBLs exposed to propofol carrier also showed lower cytotoxicity compared to propofol. No other significant differences were observed. Therefore, the documented effects of these anesthetics in vivo may be caused by an indirect mechanism. The lipid emulsion's significant decrease in PBL cytotoxicity may have implications for critically ill patients on total parenteral nutrition.


Assuntos
Anestésicos/farmacologia , Ketamina/farmacologia , Linfócitos/efeitos dos fármacos , Pregnanodionas/farmacologia , Propofol/farmacologia , Animais , Dexametasona/farmacologia , Cães
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