RESUMO
Fracture healing is a well-orchestrated and coordinated process and begins with the inflammatory stage involving the infiltration of immune cells and the release of cytokines, including tumour necrosis factor-alpha (TNF-α), interleukin-6 (IL-6) and interleukin-10 (IL-10). Low-magnitude high-frequency vibration (LMHFV) stimulation is effective in promoting fracture healing. The study hypothesis was that the innate immune response was impaired in osteoporotic fracture and LMHFV could positively modulate it. 9-month-old ovariectomy (OVX)-induced osteoporotic rats were randomised into sham (SHAM), OVX control (OVX), OVX-vibration (OVX-VT) or OVX vibration plus administration of COX-2 specific non-steroid anti-inflammatory drugs (OVX-VT-NSAID). LMHFV (35 Hz, 0.3 g) was given 20 min/d and 5 d/week to the treatment groups. Healing and innate immune response were evaluated by weekly radiographs, endpoint micro-computed tomography (µCT), enzyme-linked immunosorbent assay (ELISA) and histomorphometry at weeks 1, 2, 4 and 8 post-treatment. Results showed that OVX slightly elevated systemic inflammation but impaired the innate immune response locally at the fracture site, with significantly lower expressions of TNF-α and IL-6 but higher IL-10 expression during the early stage of healing. LMHFV was effective in accelerating the delayed fracture healing in OVX bones by partly restoring the impaired innate immune response at the fracture site, accompanied by promoted progression of macrophage polarisation from M1 (pro-inflammatory) to M2 (anti-inflammatory) phenotype. In conclusion, vibration treatment could positively modulate the impaired innate immune response and promote macrophage polarisation in osteoporotic-fracture healing.
Assuntos
Consolidação da Fratura , Macrófagos/citologia , Fraturas por Osteoporose/terapia , Vibração/uso terapêutico , Animais , Diferenciação Celular , Estrogênios/deficiência , Feminino , Imunidade Inata , Interleucina-10/genética , Interleucina-10/metabolismo , Macrófagos/metabolismo , Fraturas por Osteoporose/etiologia , Ratos , Ratos Sprague-DawleyRESUMO
Verticillium wilt caused by soil borne fungus Verticillium dahliae could significantly reduce cotton yield. The Ve1 homologous gene Gbvdr3 is resistant to Verticillium wilt. In order to understand of the function of the promoter Gbvdr3 in Gossypium barbadense, the promoter region of the receptor-like gene Gbvdr3 was obtained by genome walking, and the cis-element in the promoter was identified using the PLACE software in this study. The sequence analysis showed that the promoter contained elements related to stress resistance and light regulation. The cloned promoter was fused to the GUS reporter gene and transformed into Arabidopsis. GUS expression was specifically detected in roots, flowers, and seeds, suggesting that the expression of Gbvdr3 is tissue-specific. Separation and characterization analysis of the promoter of Gbvdr3 provides a platform for further research and application of this gene. Thorough understanding of the function of the Gbvdr3 promoter is important for better understanding of Gbvdr3 function. These results indicated that the promoter of Gbvdr3 was a tissue-specific promoter.
Assuntos
Resistência à Doença/genética , Gossypium/genética , Doenças das Plantas/genética , Proteínas de Plantas/genética , Arabidopsis/genética , Arabidopsis/crescimento & desenvolvimento , Clonagem Molecular , Regulação da Expressão Gênica de Plantas , Genoma de Planta , Gossypium/crescimento & desenvolvimento , Gossypium/virologia , Doenças das Plantas/virologia , Proteínas de Plantas/biossíntese , Raízes de Plantas , Plantas Geneticamente Modificadas/genética , Regiões Promotoras Genéticas/genética , Microbiologia do Solo , Têxteis , Verticillium/genética , Verticillium/patogenicidadeRESUMO
Hepatic progenitor cells (HPCs) are a potential cell source for liver cell transplantation but do not function like mature liver cells. We sought an effective and reliable method to induce HPC maturation. An immortalized HP14.5 albumin promoter-driven Gaussian luciferase (ALB-GLuc) cell line was established from HPCs isolated from fetal mouse liver of post coitus day 14.5 mice to investigate the effect of induction factors on ALB promoter. HP14.5 parental cells were cultured in DMEM with different combinations of 2% horse serum (HS), 0.1 µM dexamethasone (DEX), 10 ng/mL hepatic growth factor (HGF), and/or 20 ng/mL fibroblast growth factor 4 (FGF4). Trypan blue and crystal violet staining were used to assess cell proliferation with different induction conditions. Expression of hepatic markers was measured by semi-quantitative RT-PCR, Western blot, and immunofluorescence. Glycogen storage and metabolism were detected by periodic acid-Schiff and indocyanine green (ICG) staining. GLuc activity indicated ALB expression. The combination of 2% HS+0.1 µM Dex+10 ng/mL HGF+20 ng/mL FGF4 induced the highest ALB-GLuc activity. Cell proliferation decreased in 2% HS but increased by adding FGF4. Upon induction, and consistent with hepatocyte development, DLK, AFP, and CK19 expression decreased, while ALB, CK18, and UGT1A expression increased. The maturity markers tyrosine aminotransferase and apolipoprotein B were detected at days 3 and 6 post-induction, respectively. ICG uptake and glycogen synthesis were detectable at day 6 and increased over time. Therefore, we demonstrated that HPCs were induced to differentiate into functional mature hepatocytes in vitro, suggesting that factor-treated HPCs may be further explored as a means of liver cell transplantation.
Assuntos
Diferenciação Celular/efeitos dos fármacos , Embrião de Mamíferos/efeitos dos fármacos , Hepatócitos/citologia , Fígado/citologia , Células-Tronco/efeitos dos fármacos , Animais , Antígenos de Diferenciação/análise , Apolipoproteína B-100 , Apolipoproteínas B/isolamento & purificação , Proliferação de Células , Dexametasona/administração & dosagem , Fatores de Crescimento de Fibroblastos/administração & dosagem , Violeta Genciana , Glicogênio/metabolismo , Fator de Crescimento de Hepatócito/administração & dosagem , Verde de Indocianina/farmacocinética , Camundongos , Cultura Primária de Células/métodos , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Células-Tronco/citologia , Azul Tripano , Tirosina Transaminase/isolamento & purificaçãoRESUMO
Hepatic progenitor cells (HPCs) are a potential cell source for liver cell transplantation but do not function like mature liver cells. We sought an effective and reliable method to induce HPC maturation. An immortalized HP14.5 albumin promoter-driven Gaussian luciferase (ALB-GLuc) cell line was established from HPCs isolated from fetal mouse liver of post coitus day 14.5 mice to investigate the effect of induction factors on ALB promoter. HP14.5 parental cells were cultured in DMEM with different combinations of 2% horse serum (HS), 0.1 µM dexamethasone (DEX), 10 ng/mL hepatic growth factor (HGF), and/or 20 ng/mL fibroblast growth factor 4 (FGF4). Trypan blue and crystal violet staining were used to assess cell proliferation with different induction conditions. Expression of hepatic markers was measured by semi-quantitative RT-PCR, Western blot, and immunofluorescence. Glycogen storage and metabolism were detected by periodic acid-Schiff and indocyanine green (ICG) staining. GLuc activity indicated ALB expression. The combination of 2% HS+0.1 µM Dex+10 ng/mL HGF+20 ng/mL FGF4 induced the highest ALB-GLuc activity. Cell proliferation decreased in 2% HS but increased by adding FGF4. Upon induction, and consistent with hepatocyte development, DLK, AFP, and CK19 expression decreased, while ALB, CK18, and UGT1A expression increased. The maturity markers tyrosine aminotransferase and apolipoprotein B were detected at days 3 and 6 post-induction, respectively. ICG uptake and glycogen synthesis were detectable at day 6 and increased over time. Therefore, we demonstrated that HPCs were induced to differentiate into functional mature hepatocytes in vitro, suggesting that factor-treated HPCs may be further explored as a means of liver cell transplantation.
Assuntos
Animais , Camundongos , Diferenciação Celular/efeitos dos fármacos , Embrião de Mamíferos/efeitos dos fármacos , Hepatócitos/citologia , Fígado/citologia , Células-Tronco/efeitos dos fármacos , Antígenos de Diferenciação/análise , Apolipoproteínas B/isolamento & purificação , Proliferação de Células , Dexametasona/administração & dosagem , Fatores de Crescimento de Fibroblastos/administração & dosagem , Violeta Genciana , Glicogênio/metabolismo , Fator de Crescimento de Hepatócito/administração & dosagem , Verde de Indocianina/farmacocinética , Cultura Primária de Células/métodos , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Células-Tronco/citologia , Azul Tripano , Tirosina Transaminase/isolamento & purificaçãoRESUMO
The serotonin receptor 2C (HTR2C) gene has been shown to play a pivotal role in major depression. We examined the association between post-stroke depression (PSD) and polymorphism in HTR2C. A cohort of 223 patients with acute lacunar stroke admitted to the stroke unit of a university-affiliated regional hospital in Hong Kong was recruited. Three months after the onset of the index stroke, a research assistant administered the locally validated 15-item Geriatric Depression Scale. PSD was defined as a geriatric depression scale score of 7 or above. Possible confounding factors, including previous history of stroke, severity of stroke, level of social support, and recent life events, were investigated. All patients were genotyped for polymorphisms of HTR2C. Separate analyses were performed for males and females. Sixty-one patients were found to have PSD. There were significant associations between the HTR2C gene and PSD status in the male patients, but not in the female ones. After adjusting for possible confounders, the rs12837651 T allele (odds ratio = 4.020) and the rs2192371 G allele (odds ratio = 2.866) were found to be significantly associated with PSD in males. Genetic variation in HTR2C receptors appears to be involved in the pathogenesis of PSD in Chinese males.