RESUMEN
The purpose of this study was to investigate the impact of hyaluronidase (HAase) on lymphedema using an acute mouse tail lymphedema model. Six-week-old mice served to produce acute lymphedema and were then either treated with HAase injection or used as operative controls. An additional group of unmanipulated normal mice was used for comparison. Tail volumes were measured for 23 days and histological changes examined. Western blot analysis was conducted to quantify lymphatic vessel endothelial hyaluronan receptor (LYVE)-1, tumor necrosis factor (TNF)-alpha, transforming growth factor (TGF)-beta1, podoplanin, CD 44, and vascular endothelial growth factor receptor3 (VEGFR3) expression levels. The operative control group showed an increase in thickness of the dermis and subdermis, microlymphatic dilatation, and an increase in neutrophils. In contrast, the HAase treated group exhibited alleviation of inflammation evidenced by a decline in microlymphatic dilatation and neutrophils and an overall increase in microlymphatic vessels. Western blot analysis demonstrated that TNF-alpha and TGF-beta1 expression declined but CD44 expression increased in the HAase treated group. Levels of LYVE1, podoplanin, and VEGFR3 also increased significantly in the HAase group. Our results indicate that HAase treatment in the acute mouse tail model reduced lymphedema volume possibly through degradation of HA trafficking, which reduced inflammation and fibrosis in tissues and stimulated lymphangiogenesis.
Asunto(s)
Hialuronoglucosaminidasa/administración & dosificación , Linfangiogénesis/efectos de los fármacos , Vasos Linfáticos/efectos de los fármacos , Linfedema/tratamiento farmacológico , Enfermedad Aguda , Animales , Modelos Animales de Enfermedad , Femenino , Expresión Génica , Glicoproteínas/agonistas , Glicoproteínas/genética , Glicoproteínas/metabolismo , Receptores de Hialuranos/genética , Receptores de Hialuranos/metabolismo , Inyecciones Intralesiones , Linfangiogénesis/genética , Vasos Linfáticos/metabolismo , Vasos Linfáticos/patología , Linfedema/genética , Linfedema/metabolismo , Linfedema/patología , Glicoproteínas de Membrana/agonistas , Glicoproteínas de Membrana/genética , Glicoproteínas de Membrana/metabolismo , Proteínas de Transporte de Membrana , Ratones , Neutrófilos/efectos de los fármacos , Neutrófilos/patología , Cola (estructura animal) , Factor de Crecimiento Transformador beta1/antagonistas & inhibidores , Factor de Crecimiento Transformador beta1/genética , Factor de Crecimiento Transformador beta1/metabolismo , Factor de Necrosis Tumoral alfa/antagonistas & inhibidores , Factor de Necrosis Tumoral alfa/genética , Factor de Necrosis Tumoral alfa/metabolismo , Receptor 3 de Factores de Crecimiento Endotelial Vascular/agonistas , Receptor 3 de Factores de Crecimiento Endotelial Vascular/genética , Receptor 3 de Factores de Crecimiento Endotelial Vascular/metabolismoRESUMEN
OBJECTIVES: Histone deacetylase (HDAC) is an important therapeutic target in cancer. Two of the main anticancer mechanisms of HDAC inhibitors are induction of terminal differentiation and inhibition of cell proliferation. To investigate the role of HDAC in maintenance of self-renewal and cell proliferation, we treated mesenchymal stem cells (MSCs) that originated from adipose tissue or umbilical cord blood with valproic acid (VPA) and sodium butyrate (NaBu). MATERIALS AND METHODS: Human MSCs were isolated from mammary fat tissue and cord blood. We performed MTT assay and flow cytometry-based cell cycle analysis to assess self-renewal of MSCs. In vitro differentiation assays into osteogenic, adipogenic, neurogenic and chondrogenic lineages were conducted to investigate MSC multipotency. Immunocytochemistry, Western blot and reverse transcription-polymerase chain reaction were used to interrogate molecular pathways. RESULTS: VPA and NaBu flattened the morphology of MSCs and inhibited their growth. VPA and NaBu activated the transcription of p21(CIP1/WAF1) by increasing the acetylation of histone H3 and H4 and eventually blocked the cell cycle at G2/M phase. The expression level of p16(INK4A), a cdk inhibitor that is closely related to cellular senescence, was not changed by HDAC inhibitor treatment. We performed controlled differentiation into bone, fat, cartilage and nervous tissue to elucidate the role of HDAC in the pluripotency of MSC to differentiate into functional tissues. VPA and NaBu decreased the efficiency of adipogenic, chondrogenic, and neurogenic differentiation as visualized by specific staining and reverse transcription-polymerase chain reaction. In contrast, osteogenic differentiation was elevated by HDAC inhibitor treatment. CONCLUSION: HDAC activity is essential for maintaining the self-renewal and pluripotency of MSCs.
Asunto(s)
Diferenciación Celular/efectos de los fármacos , Linaje de la Célula , Proliferación Celular/efectos de los fármacos , Inhibidores Enzimáticos/farmacología , Inhibidores de Histona Desacetilasas , Células Madre Mesenquimatosas/efectos de los fármacos , Western Blotting , Ciclo Celular/efectos de los fármacos , Inhibidor p21 de las Quinasas Dependientes de la Ciclina/metabolismo , Humanos , Inmunohistoquímica , Células Madre Mesenquimatosas/citología , Osteogénesis , Reacción en Cadena de la Polimerasa de Transcriptasa Inversa , Regulación hacia ArribaRESUMEN
BACKGROUND: In addition to apolipoprotein(a) [apo(a)] kringle 4 variable number of tandem repeat (K4-VNTR), pentanucleotide repeat polymorphism (PNRP) and C/T(+93) polymorphism [C/T(+93)] of apo(a) gene have been suggested to be related to lipoprotein(a) [Lp(a)] concentration. We studied the distribution of these genetic polymorphisms and their relationship with Lp(a) concentrations in a Korean population. METHODS: One hundred thirty-two Korean adults were examined. Lp(a) was measured with enzyme-linked immunosorbent assay (ELISA). Apo(a) K4-VNTR was measured by high-resolution SDS-agarose gel separation and ECL Western blotting method. PNRP was measured after DNA amplification. The C/T(+93) ratio was measured by a amplification refractory mutation system. RESULTS: Lp(a) was inversely correlated with K4-VNTR (r=0.732, p<0.0001), but was associated neither with any PNRP haplotype nor with C/T(+93) by multiple regression analysis, although we found a significant decrease of Lp(a) in PNRP 9/9 individuals (p<0.01). There was a strong linkage disequilibrium between 9 haplotypes of PNRP and the T haplotype of C/T(+93). CONCLUSIONS: Inverse relationship between serum Lp(a) and K4 number of apo(a) was confirmed in normal Korean adults. PNRP 9/9 genotype appeared to have a reducing effect on Lp(a), but neither 9 haplotype heterozygotes of PNRP nor the T haplotype C/T(+93) affected Lp(a) concentrations in Koreans.
Asunto(s)
Apolipoproteínas A/genética , Lipoproteína(a)/genética , Polimorfismo Genético/genética , Ensayo de Inmunoadsorción Enzimática , Femenino , Genotipo , Humanos , Corea (Geográfico) , Masculino , Persona de Mediana Edad , Análisis de RegresiónRESUMEN
Reactive oxygen species (ROS), generated by infiltrating neutrophils, are considered as an important regulator in the pathogenesis and development of pancreatitis. A hallmark of the inflammatory response is the induction of cytokine gene expression, which may be regulated by oxidant-sensitive transcription factor, nuclear factor-kappaB (NF-kappaB). Present study aims to investigate whether neutrophils primed by 4beta-phorbol 12beta-myristate 13alpha-acetate (PMA) affect the productions of H(2)O(2) and lipid peroxide (LPO), NF-kappaB activation and cytokine production in pancreatic acinar cells, and whether these alterations were inhibited by N-acetylcysteine (NAC) and superoxide dismutase (SOD). Neutrophils generated ROS by stimulation with PMA, which was inhibited by NAC and SOD. In acinar cells, PMA-primed neutrophils increased the productions of H(2)O(2), LPO, and cytokines both time and dose dependently. PMA-primed neutrophils resulted in the activation of two species of NF-kappaB dimers (a p50/p65 heterodimer and a p50 homodimer) in acinar cells. Both NAC and SOD inhibited neutrophil-induced, oxidant-mediated alterations in acinar cells. In conclusion, ROS, generated by neutrophils, activates NF-kappaB, resulting in upregulation of inflammatory cytokines in acinar cells. Antioxidants such as NAC might be useful antiinflammatory agents by inhibiting oxidant-mediated activation of NF-kappaB and decreasing cytokine production.
Asunto(s)
Acetilcisteína/farmacología , Antioxidantes/farmacología , Citocinas/genética , Regulación de la Expresión Génica/inmunología , Peroxidación de Lípido/efectos de los fármacos , FN-kappa B/antagonistas & inhibidores , Páncreas/fisiología , Animales , Células Cultivadas , Citocinas/análisis , Regulación de la Expresión Génica/efectos de los fármacos , Peróxido de Hidrógeno/metabolismo , Cinética , Masculino , Páncreas/efectos de los fármacos , Páncreas/inmunología , ARN Mensajero/genética , Ratas , Ratas Sprague-Dawley , Superóxido Dismutasa/metabolismo , Acetato de Tetradecanoilforbol/farmacología , Factores de Tiempo , Transcripción Genética/efectos de los fármacosRESUMEN
Most reports on serious MTX toxicity have focused on hepatic abnormalities, while other effects, including hematologic reactions, have not been emphasized. We experienced a case of pancytopenia secondary to MTX therapy in a patient with RA and renal insufficiency. A 67-year-old woman with a 12-year history of active seropositive RA that was a response to non-steroidal anti-inflammatory drugs, hydroxychloroquinine and intra-articular steroid injections, had been followed up and was diagnosed as early chronic renal failure in October, 1993. Recently, because of significant morning stiffness and polyarthralgia, the decision was made to institute MTX treatment. This was begun as a single oral dose of 5mg/week. After 2 doses, the patient was admitted to the hospital with general weakness. Laboratory tests showed a hemoglobin level of 7.9 g/dl, WBC count 1800/mm3 and platelet count of 64000/mm3. The serum creatinine level was 6.1 mEq/dl and the BUN level was 82 mEq/dl. Liver function test results were normal, but the serum albumin level was 2.7 g/dl. The patient subsequently developed fever and blood transfusions, granulocyte colony stimulating factor (G-CSF) and intravenous prophylactic antibiotic therapy were required. Her condition was improved. In summary, Low-dose MTX-related adverse hematologic side effects, including fatal pancytopenia, are rare but are a cause of increasing concern in patients with RA and renal insufficiency. Close monitoring of associated risk factors, particularly impaired renal function, should be mandatory for all patients who are receiving MTX therapy.