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OBJECTIVE: Pulmonary aspergillosis, which is a secondary complication of fungal pneumonia, is widely considered to have an increasing incidence and high mortality. Itraconazole (Itz) can inhibit ergosterol biosynthesis to treat pulmonary aspergillosis. Nevereless, Itz's clinical application is limited because of its poor water solubility, low oral bioavailability, and systemic hepatotoxicity. In this study, Itz-loaded nanostructured lipid carriers (Itz-NLCs) were developed to improve the in vitro permeability and bioavailability of Itz via pulmonary administration. METHODS: Itz-NLCs were prepared by the emulsification-evaporation method using oleic acid and glycerol monostearate as liquid and solid lipids, respectively. RESULTS: The Itz-NLCs were optimized with tiny particle size, uniform distribution, and excellent entrapment efficiency (EE, 97.57% ± 0.45%). A Xenopus alveolar membrane was used in the permeation study, and the cumulative permeation percentage of Itz was 10% for Itz-NLCs at 8 h, which was 2.50-fold higher than that for Itz suspensions (4%, p < .001). A rabbit pharmacokinetic investigation revealed that Itz-NLCs have an 83.05% absolute bioavailability after intratracheal instillation. CONCLUSIONS: The purpose of Itz-NLCs is to enhance the bioavailability and permeability of Itz in vitro for administration via the lungs.
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Nanoestructuras , Aspergilosis Pulmonar , Animales , Conejos , Itraconazol/farmacología , Portadores de Fármacos , Administración Oral , Lípidos , Disponibilidad Biológica , Tamaño de la PartículaRESUMEN
Multidrug resistance has dramatically compromised the effectiveness of paclitaxel (PTX). The combined application of PTX and tetrandrine (TET) is a promising avenue in drug-resistant cancer therapy. However, poor drug release and limited intracellular drug accumulation greatly impede this combinational antitumor therapy. To address this problem, we successfully developed a tunable controlled release lipid platform (PT@usNLC) for coordinated drug delivery. The drug release rate of PT@usNLC can be tuned by varying the lipid ratio, which has potential to maximize the therapeutic effects of combined drugs. The TET release rate from PT@usNLC was faster than PTX, which could restore the sensitivity of tumor cells to PTX and exert a synergistic antitumor effect. The appropriate size of PT@usNLC could effectively increase the intracellular drug accumulation. Bothin vitroandin vivostudies revealed that PT@usNLC significantly enhanced the therapeutic effect compared to conventional therapies. This study provides a new strategy for resistant ovarian cancer therapy.
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Nanopartículas , Neoplasias Ováricas , Línea Celular Tumoral , Portadores de Fármacos , Femenino , Humanos , Lípidos , Neoplasias Ováricas/tratamiento farmacológico , Paclitaxel/farmacología , Preparaciones FarmacéuticasRESUMEN
This study describes variations in tumour growth patterns which occur when changes in the routes of inoculation and mouse strain are used to introduce tumours into established murine model systems that are known to vary in location and aggression. Intraperitoneal, subcutaneous, intravenous and hydrodynamic inoculations of B16F10 cells were compared among CD-1, C57BL/6 and Balb/c mice. Most surprisingly, allogeneic tumour growth in Balb/c mice after intravenous and hydrodynamic inoculation of B16F10 cells was faster than tumour growth in the syngeneic C57BL/6 mice. These and other variations in the tumour growth patterns described here can help provide the researcher with more experimental control when planning to use the optimal tumour model for any particular study.
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Modelos Animales de Enfermedad , Melanoma Experimental/patología , Animales , Femenino , Hidrodinámica , Inyecciones Intraperitoneales , Inyecciones Intravenosas , Inyecciones Subcutáneas , Ratones Endogámicos BALB C , Ratones Endogámicos C57BL , Trasplante de Neoplasias/métodos , Especificidad de la EspecieRESUMEN
1. Ephedra water decoction (EWD) and cough tablets containing ephedra and liquorice (maxing cough tablets, MXCT) have been widely used in the treatment of asthma. In the clinic, EWD and MXCT may be prescribed with theophylline, one of the most popular antiasthmatic drugs. CYP1A2 and CYP2E1 are mainly involved in the oxidative metabolism of theophylline in human liver. Drug interactions involving the cytochrome P450 (CYP) isoforms generally are of two types: enzyme induction or enzyme inhibition. Enzyme inhibition reduces metabolism, whereas induction can increase it. 2. To evaluate the pretreatment effect of EWD and MXCT on CYP1A2 and CYP2E1, CYP1A2 and CYP2E1 activity, the protein expression and mRNA expression levels were determined. After pretreatment with EWD or MXCT, the enzyme activity, mRNA expression and protein expression of CYP1A2 were increased significantly (p < 0.05), but enzyme activity of CYP2E1 did not change compared with the control. 3. It was demonstrated that EWD or MXCT pretreatment obviously induced CYP1A2, therefore, in patients taking EWD or MXCT, possible CYP-induced drug interaction should be noted to decrease the risk of therapeutic failure or adverse effects resulting from the use of additional therapeutic agents.
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Citocromo P-450 CYP2E1/metabolismo , Citocromos/metabolismo , Ephedra/química , Glycyrrhiza/química , Hígado/efectos de los fármacos , Preparaciones de Plantas/farmacología , Animales , Antiasmáticos/farmacología , Citocromo P-450 CYP1A2 , Hígado/enzimología , Masculino , Microsomas Hepáticos/efectos de los fármacos , Microsomas Hepáticos/metabolismo , ARN Mensajero/metabolismo , Ratas , Ratas Wistar , Comprimidos , Teofilina/farmacologíaRESUMEN
The aim of this study was to design novel mixed micelles as an ophthalmic delivery system for alpha-tocopherol (TOC) to prevent its degradation and improve ocular efficacy. The nonionic polymers, Polyoxyl 15 Hydroxystearate (Solutol® HS15) and Pluronic® F127, were discovered to be the most effective agents for retaining the activity and solubilization of TOC, respectively. Prepared by a thin-film hydration method, HS15/Pluronic® F127 yielded good encapsulation percentages of TOC, with a 27.7% drug loading efficiency. Incorporation of cetalkonium chloride (CKC) into HS15/Pluronic® F127 mixed micelles made the zeta potential of the micelles +17 mV, potentially prolonging the residence time of formulations on ocular surfaces. The optimized micelle preparation remained stable when diluted in a synthetic tear solution. It is known that the antioxidant ability of TOC in typical formulations reduces to around 85% of its initial value after 1 month when stored at 4 or 25 °C under an air atmosphere, which limits ophthalmic applications to less than 1 month. However, encapsulated TOC in investigated micelles remained stable for at least 6 months when sealed with N2. Finally, the cationic micelles were well tolerated after multiple administrations in rabbits, and they improved ocular accumulation of TOC. Taken together, these data suggest that the optimized micelle preparations described in this study may be suitable drug carriers for the treatment of ocular oxidant damage.
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Antioxidantes/administración & dosificación , Córnea/efectos de los fármacos , Soluciones Oftálmicas/administración & dosificación , Oxidantes/antagonistas & inhibidores , alfa-Tocoferol/administración & dosificación , Animales , Antioxidantes/metabolismo , Córnea/metabolismo , Portadores de Fármacos/administración & dosificación , Portadores de Fármacos/metabolismo , Estabilidad de Medicamentos , Masculino , Micelas , Soluciones Oftálmicas/metabolismo , Oxidantes/metabolismo , Conejos , alfa-Tocoferol/metabolismoRESUMEN
Although the development of multi-disciplinary management has improved the survival of colorectal cancer (CRC), the prognosis of metastatic CRC patients remains poor. Accumulating evidence has demonstrated that immunotherapy with cancer vaccines and adoptive T cell transfusions may improve outcomes as an adjuvant to current standard CRC treatment. In this phase I/II study, 71 CRC patients who underwent radical surgery (stage I-III, n = 46) or palliative surgery (stage IV with non-resectable synchronous metastases, n = 25) were included. In the first part of this study, sentinel lymph nodes (SLNs) were intraoperatively identified in 55 patients (46 with stage I-III CRC and 9 with stage IV CRC). SLN-T lymphocytes were expanded ex vivo for a median of 28.5 days (range 23-33 days). Thereafter, a median of 153 × 10(6) cells (range 20.7-639.0 × 10(6)) were transfused. No treatment-related toxicity was observed. In the second part of this study, the stage IV patients were routinely followed. The 24-month survival rate of the SLN-T lymphocyte group was significantly higher than that of the control group: 55.6 versus 17.5% (p = 0.02). The median overall survival of the SLN-T lymphocyte and control groups was 28 and 14 months, respectively. Our study showed that adjuvant SLN-T lymphocyte immunotherapy is feasible and safe for postoperative CRC patients. Additionally, this therapy may improve the long-term survival of metastatic CRC. Further investigation of the clinical efficacy and anti-tumor immunity is warranted.
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Neoplasias Colorrectales/inmunología , Neoplasias Colorrectales/terapia , Inmunoterapia Adoptiva/métodos , Inmunoterapia/métodos , Ganglios Linfáticos/patología , Linfocitos T/inmunología , Linfocitos T/trasplante , Adyuvantes Inmunológicos , Anciano , Neoplasias Colorrectales/patología , Femenino , Humanos , Masculino , Persona de Mediana Edad , Pronóstico , Linfocitos T/patologíaRESUMEN
BACKGROUND: Vaginal inflammation is a prevalent gynecological condition. If left untreated, it can potentially spread to the urinary and reproductive systems. METHODS: In this study, we propose a resveratrol-loaded microemulsion-based thermosensitive hydrogel (Res-Me-Tsgel) and compare it with a chitosan hydrogel-based Res-Me-Cogel. We characterized the different characters of Res-Me-Tsgel. The safety of Res-Me-Tsgel was also evaluated in vitro and in vivo. Finally, we measured the retention of Res in the vagina after drug administration. RESULTS: The Res-Me-Tsgel we prepared is a transparent liquid solution at room temperature that rapidly forms a gel at 37oC. Compared to Res solution and Res-Me, both Res-Me-Cogel and Res-Me-Tsgel demonstrate superior sustained release properties. Both in vitro and in vivo studies confirm the excellent biosafety profile of Res-Me-Cogel and Res-Me-Tsgel. Vaginal administration of these formulations in rats results in prolonged retention of resveratrol within the vagina. Notably, due to its improved flow into vaginal folds after administration, the retention of Resveratrol was approximately three times higher for the Res-Me-Tsgel group compared to the Res-Me-Cogel group at 24 h post-administration. Overall, these findings highlight the potential application of Res-Me-Tsgel as an effective means for vaginal inflammation. CONCLUSIONS: We developed a novel micromulsion based thermosensitive hydrogel for the delivery of Res. The sustained release of Res and favorable vaginal retention from Res-Me-Tsgel make them promise as a potential candidate for local intravaginal therapy.
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Hidrogeles , Inflamación , Femenino , Ratas , Animales , Resveratrol/farmacología , Preparaciones de Acción Retardada , Administración Tópica , Inflamación/tratamiento farmacológicoRESUMEN
Multidrug resistance (MDR) is a major cause of failure in cancer chemotherapy. Tocopheryl polyethylene glycol 1000 succinate (TPGS) has been extensively explored for the treatment of MDR in cancer because of its ability to inhibit P-glycoprotein. Here, we have established multifunctional nanoparticles (MFNPs) using a single-molecule modification of TPGS, which can deliver a hydrophobic drug, paclitaxel (PTX), and a hydrophilic drug, fluorouracil (5-FU), and overcome MDR in cancer. Our data indicated that, when delivered into a PTX-resistant cell line using MFNPs, the combination of PTX and 5-FU was more cytotoxic than each agent individually.
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Antineoplásicos/química , Antineoplásicos/farmacología , Resistencia a Múltiples Medicamentos , Resistencia a Antineoplásicos , Nanopartículas/química , Neoplasias/tratamiento farmacológico , Subfamilia B de Transportador de Casetes de Unión a ATP/química , Miembro 1 de la Subfamilia B de Casetes de Unión a ATP/metabolismo , Línea Celular Tumoral , Diseño de Fármacos , Citometría de Flujo , Fluorouracilo/farmacología , Humanos , Hidrólisis , Modelos Químicos , Paclitaxel/farmacología , Polietilenglicoles/farmacología , Rodamina 123/farmacología , Vitamina E/análogos & derivados , Vitamina E/farmacologíaRESUMEN
Multidrug resistance (MDR) is a major barrier to the chemotherapy treatment of many cancers. However, some nonionic surfactants, for example, Brij, have been shown to restore the sensitivity of MDR cells to such drugs. The aim of this study was to explore the reversal effect of Brij on MDR tumor cells and elucidate its potential mechanism. Our data indicate that the structure of Brij surfactants plays an important role in overcoming MDR in cancer, that is, modified hydrophilic-lipophilic balance (MHLB, the ratio of the number (n) of hydrophilic repeating units of ethylene oxide (EO) to the number (m) of carbons in the hydrophobic tail (CH(2))). Cell viability of cells treated with paclitaxel (PTX) nanocrystals (NCs) formulated with Brij showed positive correlations with MHLB (R(2) = 0.8195); the higher the ratio of Brij to PTX in NCs, the higher cytotoxicity induced by the PTX NCs. Significant increases in intracellular accumulation of (3)H-PTX (P-gp substrate) were observed in an MDR cell line (H460/taxR cells) treated with Brij 78 (MHLB = 1.11) and Brij 97 (MHLB = 0.6). After treatments with Brij 78 and Brij 97, the levels of intracellular ATP were decreased and verapamil-induced ATPase activities of P-gp were inhibited in multidrug resistant cells. The responses of the cells to Brij 78 and Brij 97 in ATP depletion studies correlated with the cell viability induced by PTX/Brij NCs and intracellular accumulation of (3)H-PTX. Brij 78 and Brij 97 could not alter the levels of P-gp expression detected by Western blotting. These findings may provide some insight into the likelihood of further development of more potent P-gp inhibitors for the treatment of MDR in cancer.
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Miembro 1 de la Subfamilia B de Casetes de Unión a ATP/antagonistas & inhibidores , Adenosina Trifosfatasas/antagonistas & inhibidores , Adenosina Trifosfato/metabolismo , Aceites de Plantas/farmacología , Polietilenglicoles/farmacología , Antineoplásicos Fitogénicos/farmacología , Transporte Biológico/efectos de los fármacos , Bloqueadores de los Canales de Calcio/farmacología , Línea Celular Tumoral , Supervivencia Celular/efectos de los fármacos , Resistencia a Múltiples Medicamentos , Resistencia a Antineoplásicos , Humanos , Interacciones Hidrofóbicas e Hidrofílicas , Neoplasias Pulmonares/tratamiento farmacológico , Nanopartículas , Paclitaxel/farmacología , Polidocanol , Tensoactivos/farmacología , Verapamilo/farmacologíaRESUMEN
Patients with sepsis face high mortality rates and a bleak prognosis, prompting the need for advanced therapeutic interventions. A male patient diagnosed with moderately low-differentiated squamous cell carcinoma received diverse treatments, including radiotherapy, chemotherapy, immunotherapy, and targeted therapy to inhibit angiogenesis. Subsequently, he developed sepsis after comprehensive treatment, and conventional antibiotic combinations proved ineffective in combating the infection. As an experimental approach, allogeneic natural killer (NK) cell infusion was administered. Following the NK cell infusion, the patient regained consciousness, and laboratory analyses showed reduced infection-related markers, suppressed serum inflammatory cytokines, and elevated anti-tumor cytokines. However, the therapeutic effect only lasted 2-3 days. In vitro investigations demonstrated that the allogeneic NK cell product reduced interleukin-6 levels in the patient's serum. Moreover, subsequent co-cultivation of the NK cell product with the patient's serum resulted in a decrease in the proportion of cytotoxic subpopulations of NK cells and a downregulation of the expression of NK-mediated killing molecules. In conclusion, adoptive transfusion of allogeneic NK cells may improve sepsis symptoms in patients with tumor-related sepsis. In vitro co-culture tests hold promise in providing predictive biomarkers for treatment effectiveness.
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The detection of colorectal cancer (CRC) lymph node (LN) metastases significantly influences treatment choices, yet identifying them in samples is time-consuming and error-prone. To enhance efficiency, we have established a LN metastasis detection method utilizing triple-parameter flow cytometry (tFCM) and have conducted a comparative assessment of its accuracy and cost-effectiveness in contrast to conventional pathological examinations. This technique utilized biomarkers cytokeratin 20 (CK20), epithelial cell adhesion molecules (EpCAM), and Pan-CK. tFCM's sensitivity was validated by analyzing known cell line concentrations (SW480 and SW620) in peripheral blood mononuclear cells (PBMCs), with CK20, EpCAM, and Pan-CK showing significant expression in CRC cell lines but not in PBMCs. A strong linear correlation was observed in the mixed leukocyte environment (R 2 = 0.9988). Subsequently, tFCM and pathological sections were employed to analyze LNs from CRC patients, enabling comparison of detection accuracy. Within the 36 LNs studied, tFCM successfully identified tumor cells with varying metastasis degrees, including micro-metastasis and isolated tumor cell clusters. Notably, relying solely on pathological sections led to a potential 25% misdiagnosis rate for LNs. In contrast, tFCM effectively minimized this risk. In summary, compared to traditional pathological sections, tFCM is a more advantageous method for detecting nodal metastasis in CRC patients, offering a more precise prognosis for these patients.
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Ephedra water decoction (EWD) and cough tablets containing ephedra and liquorice (maxing cough tablets, MXCT) have been used widely in the treatment of asthma. In the clinic, EWD and MXCT may be prescribed with theophylline, one of the most popular antiasthmatic drugs and a typical substrate of cytochrome P450 (CYP) 1A2. So in the present study the potential effects of EWD and MXCT on CYP1A2 activity and the pharmacokinetics of theophylline in rats were evaluated. In the in vivo CYP1A2 activity research, the rats were given oral caffeine (10 mg/kg) after a 14 day pretreatment with EWD (18 g/kg) and MXCT (0.1, 0.2 or 0.4 g/kg). Then the CYP 1A2 activity was expressed by using the caffeine metabolic ratio (CMR). The results showed that the CMR increased markedly compared with the control groups. In the pharmacokinetics experiment, the rats were given oral theophylline (10 mg/kg) after a 14 day pretreatment with EWD (18 g/kg) and MXCT (0.2 g/kg). The results showed that the AUC(0-24 h) and C(max) of theophylline were reduced markedly compared with the control groups. These results demonstrated that EWD or MXCT pretreatment obviously induced CYP1A2 activity, therefore, speeding up the metabolism of theophylline. The concomitant use of EWD or MXCT may decrease the effect of theophylline in rats.
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Citocromo P-450 CYP1A2/química , Ephedra/química , Glycyrrhiza/química , Teofilina/farmacocinética , Animales , Cafeína/administración & dosificación , Inhibidores del Citocromo P-450 CYP1A2 , Activación Enzimática , Inhibidores Enzimáticos/química , Inhibidores Enzimáticos/farmacología , Hígado/química , Hígado/efectos de los fármacos , Hígado/enzimología , Masculino , Distribución Aleatoria , Ratas , Ratas Wistar , Comprimidos/química , Teofilina/administración & dosificación , Teofilina/química , Factores de Tiempo , Agua/químicaRESUMEN
Primary cytoreductive surgery with platinum-taxane-based chemotherapy is the standard treatment for ovarian cancer (OC) patients; however, resistance to chemotherapy is a contributing factor to OC mortality. Paclitaxel (PTX), the most widely used taxane, has become the first-line drug against OC. The molecular mechanism of PTX resistance is different from that of platinum-based agents and is still not completely elucidated. Our previous study showed that glucose-regulated protein 78 (GRP78) is involved in the resistance of OC cells to PTX. However, little is known regarding endogenous inhibitors of this gene. MicroRNAs (miRNAs) play critical roles in the regulation of gene expression; therefore, we sought to identify miRNA(s) with potential to target GRP78 under the hypothesis that miRNA(s) could serve as potential therapeutic targets. Here, we show that miR-181c, predicted to target GRP78, was downregulated in PTX-resistant OC cells and tissues. MiR-181c downregulated GRP78 expression and induced apoptosis by directly targeting its 3'-untranslated region (UTR). Overexpression of miR-181c sensitized resistant OC to PTX by inhibiting the PI3K/Akt pathway in vitro and in vivo. Taken together, our findings indicate that the delivery of miR-181c can efficiently suppress GRP78 expression and GRP78-mediated PTX resistance in OC and suggest that this strategy has therapeutic potential.
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MicroARNs , Neoplasias Ováricas , Regiones no Traducidas 3' , Carcinoma Epitelial de Ovario/genética , Línea Celular Tumoral , Resistencia a Antineoplásicos/genética , Chaperón BiP del Retículo Endoplásmico/metabolismo , Femenino , Regulación Neoplásica de la Expresión Génica , Humanos , MicroARNs/genética , MicroARNs/metabolismo , Neoplasias Ováricas/tratamiento farmacológico , Neoplasias Ováricas/genética , Neoplasias Ováricas/metabolismo , Paclitaxel/farmacología , Paclitaxel/uso terapéutico , Fosfatidilinositol 3-Quinasas/genética , Fosfatidilinositol 3-Quinasas/metabolismo , Proteínas Proto-Oncogénicas c-akt/genética , Proteínas Proto-Oncogénicas c-akt/metabolismoRESUMEN
BACKGROUND: Pulmonary arterial hypertension (PAH) is characterized by abnormal proliferation of vascular endothelial and smooth muscle cells and causes occlusion of pulmonary arterioles that eventually results in right heart failure and death. The platelet-derived growth factor (PDGF) plays a prominent role in abnormal remodeling of pulmonary resistance vessels. Imatinib mesylate (IM), a PDGF-receptor tyrosine kinase inhibitor, was able to ameliorate PAH by reversing pulmonary vascular remodeling. METHODS: In the present study, IM-loaded liposomes (IM-LPs) were developed and administered via the pulmonary route to delay the drug release and improve patient compliance for the treatment of PAH. The IM-LPs were prepared by the transmembrane gradient method with the spherical vesicles. The compatibility of the IM-LPs was studied by determining the viability of pulmonary arterial smooth muscle cells (PASMCs). Particle uptake by rat PASMCs was evaluated by incubating the particles with rat PASMCs. Pharmacokinetic studies were performed in male SD rats. RESULTS: The IM-LPs showed an average size of 101.6 ± 50.80 nm with a zeta potential value of 19.66 ± 0.55 mV, a PDI of 0.250 and 81.96% ± 0.98% drug entrapment efficiency, meanwhile displayed a sustained release profile. Liposomes obviously increased intracellular accumulation of Rhodamine B by PASMCs using the fluorescence microscopic. Following intratracheal administration to rats, IM-LPs not only extended the half-life of IM, but also prolonged retention of IM compared with plain IM solution after intratracheal and intravenous administration. CONCLUSION: The study show potential applications of the LPs for pulmonary delivery of IM and the method for the development of LPs in sustained release of IM for better therapeutic outcomes. Conclusively, the prepared IM-LPs were well designed in nanosized ranges and may be a promising formulation for pulmonary delivery of IM.
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Sistemas de Liberación de Medicamentos , Mesilato de Imatinib/farmacocinética , Pulmón/efectos de los fármacos , Animales , Proliferación Celular/efectos de los fármacos , Supervivencia Celular/efectos de los fármacos , Liberación de Fármacos , Mesilato de Imatinib/sangre , Mesilato de Imatinib/farmacología , Liposomas , Masculino , Miocitos del Músculo Liso/efectos de los fármacos , Miocitos del Músculo Liso/metabolismo , Arteria Pulmonar/citología , Ratas Sprague-DawleyRESUMEN
Lymph node metastasis indicates a poor prognosis in colorectal cancer. To better understand the underlying mechanisms of lymph node metastasis, we analyzed transcriptome characteristics of the pre-metastatic lymph node, a putative microenvironment favorable for the seeding and proliferation of cancer cells. Thus, we tried to compare and elucidate the transcriptional and immune characteristics of sentinel lymph nodes (SNs) with matched non-sentinel lymph nodes (NSNs) in colorectal cancer patients. In this study, a total of 38 pairs of SNs and NSNs were collected, in which 26 pairs of non-metastatic lymph nodes were subjected to RNA-seq and bioinformatics analysis for the gene expression profiles. There were 16 differentially expressed genes between SNs and NSNs being identified, including 9 upregulated and 7 downregulated genes in SN. Gene Ontology (GO) classification analysis revealed that the differentially expressed genes were mainly involved in leukocyte differentiation, chemokine secretion, and immune system regulation. In the meantime, gene set enrichment analysis (GSEA) showed that immune-related signaling pathways, such as transforming growth factor beta (TGF-ß) signaling and tumor necrosis factor alpha (TNF-α)/nuclear factor kappa-light-chain-enhancer of activated B cells (NF-κB) signaling, were enriched in NSN, while cell proliferation-related signaling pathways were enriched in SN, including MYC signaling and G2M checkpoint signaling. We further identified SIGLEC15 as a top upregulated gene in SN. However, RNAscope assay showed that SIGLEC15 was not largely co-expressed with M2 macrophage marker CD163. We then selected eight pairs of lymph nodes for further cytological studies. Flow cytometry analysis revealed that Siglec-15 was expressed on all myeloid cell subsets. The relative expression of SEGLEC15 (SN/NSN) was correlated with the microsatellite instability (MSI) status in colorectal cancer patients. Further studies found that small interfering ribonucleic acid (siRNA)-mediated silencing of SLGLEC15 can enhance the anti-tumor function of T cells, as indicated by cytokine release analysis. In conclusion, we presented here a first report on the gene expression profiling of the pre-metastatic lymph node in colorectal cancer. The findings in this study suggest that SIGLEC15 plays an important role in SN immunosuppression. SEGLEC15 silencing could be a therapeutic strategy for restoring T cell function in tumor SNs.
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PURPOSE: This study is aimed at immunologically characterizing sentinel lymph nodes (SNs) in colorectal cancer (CRC) patients and identifying changes in immunological phenotype and function of SNs isolated from the tumor immunosuppressive microenvironment. METHODS: A total of 53 pairs of matched SNs and non-SNs (NSNs) were collected by using a lymph node tracer dye. Flow cytometry was performed to detect the immunophenotype of T cells as well as the expression of activation and inhibitory markers. Differential expression and distribution of characteristic immune cell markers were analyzed by multiplex immunohistochemistry (mIHC). Transcriptomics analysis was conducted to compare the differences in the expression of immune-related genes among lymph nodes. The ex vivo culture of lymph nodes was carried out to examine changes in immunological phenotypes and functions. RESULTS: Compared with NSNs, SNs harbored a significantly higher percentage of regulatory T cells (Tregs) but a lower proportion of MoMDSCs. As indicated in the mIHC assays, Tregs, T follicular helper (Tfh) cells, and M2 macrophages were mainly distributed in cortical areas, germinal centers, and subcapsular sinus areas, respectively, while significantly higher numbers of Tregs and Tfh cells were detected in SNs as compared to NSNs. Moreover, GSEA revealed that T cell activation genes and CD8+ T cell exhaustion-related genes are enriched in SNs and NSNs, respectively. The ex vivo culture led to an increase in the proportion of CD4+ cells, while activating T cells in SNs. In addition, SNs displayed a higher increase in the expression of cytokines IFN-γ, TNF-α, and sFas than NSNs. CONCLUSION: SNs are shown to be in an immune active state in vivo, while highly expressing inhibitory cytokines and suppressive markers. The ex vivo culture enhanced antitumor immunological function of SN-T cells, providing a starting material for adoptive cell therapy for CRC.
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Neoplasias Colorrectales/inmunología , Inmunoterapia Adoptiva/métodos , Ganglio Linfático Centinela/inmunología , Linfocitos T Reguladores/inmunología , Adulto , Anciano , Anciano de 80 o más Años , Neoplasias Colorrectales/diagnóstico , Neoplasias Colorrectales/patología , Neoplasias Colorrectales/terapia , Femenino , Citometría de Flujo , Perfilación de la Expresión Génica , Humanos , Inmunofenotipificación , Masculino , Persona de Mediana Edad , Estadificación de Neoplasias , Ganglio Linfático Centinela/citología , Ganglio Linfático Centinela/patología , Biopsia del Ganglio Linfático Centinela , Linfocitos T Reguladores/trasplante , Microambiente Tumoral/inmunologíaRESUMEN
BACKGROUND: To reveal the underling molecular mechanism in brain damage induced by chronic fluorosis, the neurotoxicity and its correlation were investigated by transcriptomics and proteomics. METHODS: Sprague-Dawley rats were treated with fluoride at different concentrations (0, 5, 50 and 100â¯ppm, prepared by NaF) for 3 months. Spatial learning and memory were evaluated by Morris water maze test; neuronal morphological change in the hippocampus was observed using Nissl staining; and the level of oxidative stress including reactive oxygen species (ROS), malondialdehyde (MDA) and superoxide dismutase (SOD) were detected by biological methods. The high-throughput transcriptome sequencing (RNA-Seq) and tandem mass tag (TMT) proteomic sequencing were performed to detect the expression of differentially expressed genes and proteins, respectively. RESULTS: The results showed that compared with control group, rats exposed to high-dose fluoride exhibited declined abilities of learning and memory, decreased SOD activity and increased ROS and MDA levels, with lighter colored Nissl bodies. A total of 28 important differentially expressed genes (DEGs) were screened out by transcriptomics. Then, functional enrichment analyses showed that upregulated proteins enriched in cellular transport, while downregulated proteins enriched in synapse-related pathways. Thirteen corresponding DEGs and DAPs (cor-DEGs-DAPs) were identified by differential expressions selected with positively correlated genes/proteins, most of which were related to neurodegenerative changes and oxidative stress response. CONCLUSION: These results provide new omics evidence that rats chronically exposed to high-dose fluoride can induce neurotoxicity in the brains through changes in the cholinergic pathway and oxidative stress.
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Colinérgicos/toxicidad , Fluoruros/toxicidad , Hipocampo/efectos de los fármacos , Proteómica , Animales , Colinérgicos/administración & dosificación , Relación Dosis-Respuesta a Droga , Femenino , Fluoruros/administración & dosificación , Hipocampo/metabolismo , Masculino , Estrés Oxidativo/efectos de los fármacos , Estrés Oxidativo/genética , Ratas , Ratas Sprague-Dawley , TranscriptomaRESUMEN
A sensitive and specific method based on liquid chromatography-tandem mass spectrometry using electrospray ionization (LC-ESI-MS/MS) has been developed for the determination of Schisandrin and Schisandrin B in rat plasma. A 100 microL plasma sample was extracted by methyl tert-butyl ether after spiking the samples with nimodipine (internal standard) and performed on an XTerra(R)MS-C(18) column (150 mm x 2.1 mm, 3.5 mum) with the mobile phase of acetonitrile-water-formic acid (80:20:0.2, v/v) at a flow rate of 0.2 mL/min in a run time of 8.5 min. The lower limit of quantification of the method was 40 ng/mL for Schisandrin and 20 ng/mL for Schisandrin B. The method showed reproducibility with intra-day and inter-day precision of less than 13.8% RSD, as well as accuracy, with inter- and intra-assay accuracies between 93.5 and 107.2%. Finally, the LC-ESI-MS/MS method was successfully applied to study the pharmacokinetics of Schisandrin and Schisandrin B in rats after administration of Wurenchun commercial formulations to rats.
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Cromatografía Liquida/métodos , Ciclooctanos/sangre , Medicamentos Herbarios Chinos/administración & dosificación , Lignanos/sangre , Compuestos Policíclicos/sangre , Espectrometría de Masa por Ionización de Electrospray/métodos , Animales , Masculino , Ratas , Ratas Sprague-Dawley , Espectrometría de Masas en Tándem/métodosRESUMEN
BACKGROUND: Wurenchun is the alcohol extract from Fructus Schisandrae Chinensis, which has good efficiency in lowering abnormal serum glutamic pyruvic transaminase (SGPT) level of patients suffering from acute or chronic hepatitis. The main purpose of this work is to prepare self-emulsifying drug delivery systems (SEDDS) for enhancing the solubility, dissolution rate, and oral bioavailability of poorly water-soluble traditional Chinese medicines, Wurenchun. METHODS: Pseudoternary phase diagrams were used to evaluate the efficient self-emulsification domains, and particle size distributions of resultant emulsions were determined. The dissolution test was performed according to the second method of Chinese Pharmacopoeia dissolution procedure. The pharmacokinetic study in rats for the optimized formulation was performed and compared to commercial capsules. RESULTS: The optimized formulation for bioavailability assessment consisted of 20% oleic acid, 65% Tween 20, and 15% Transcutol P. The mean droplet size distribution of the optimized SEDDS was approximately 240 nm when diluted with 1000-fold volume of the distilled water. The in vitro dissolution rates of the active components of Wurenchun SEDDS were significantly higher than those of the commercial capsules. SEDDS have significantly increased the C(max) and area under the curve) (AUC) of Wurenchun compared to reference capsules (P < 0.05). And the relative bioavailability of SEDDS for schisandrin and schisandrin B was 292.2% and 205.8% compared to the commercial capsules, respectively. CONCLUSION: The results suggest the potential use of SEDDS to improve oral absorption of the sparingly soluble drugs or traditional Chinese medicine, such as Wurenchun.
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Sistemas de Liberación de Medicamentos , Excipientes/química , Extractos Vegetales/farmacocinética , Schisandra/química , Administración Oral , Animales , Área Bajo la Curva , Disponibilidad Biológica , Ciclooctanos/farmacocinética , Medicamentos Herbarios Chinos/administración & dosificación , Medicamentos Herbarios Chinos/farmacocinética , Emulsiones , Frutas , Lignanos/farmacocinética , Masculino , Tamaño de la Partícula , Extractos Vegetales/administración & dosificación , Compuestos Policíclicos/farmacocinética , Ratas , Ratas Sprague-Dawley , SolubilidadRESUMEN
In this study, we investigated the feasibility of dipalmitoylphosphatidylcholine-coated lipid nanoparticles (DPPC-LNs) as a carrier for preferential accumulation into lungs of Resveratrol (Res), a potentially promising drug for the treatment of pulmonary arterial hypertension (PAH). Res-loaded DPPC-LNs were prepared following a thin film hydration-ultrasonic dispersion technique using glyceryl monostearate as lipid core. DPPC can reduce the interactions between nanoparticles and pulmonary surfactant. The optimal formulation was prepared and characterized for physicochemical properties, storage stability and in vitro release profiles. The optimal formulation was evaluated for uptake by pulmonary arterial smooth muscle cells (PASMCs) using fluorescence microscopy. The efficacy of Res-loaded DPPC-LNs in reducing hyperplasia was tested in 5-HT induced proliferated PASMCs. The drug absorption profiles upon intratracheal administration were monitored in healthy rats. Optimized spherical DPPC-LNs - with mean size of 123.7 nm, zeta potential of -19.4 mV and entrapment efficiency of 94.40% - exhibited an 80% cumulative drug release over 48 h. Fluorescence microscopic study revealed an time-dependent enhancement of cellular uptake of Rh123-labeled DPPC-LNs by PASMCs. PASMC proliferation induced by 5-HT was significantly inhibited by Res-loaded DPPC-LNs. Optimized DPPC-LNs appeared to be safe when incubated with PASMCs. Besides, plasma and lung tissue data analysis indicated higher value of accumulation after intratracheal administration of Res-loaded DPPC-LNs in comparison with the intravenously dosed Res solution, indicating longer retention of Res in the lungs and their slower entry to the systemic blood circulation. DPPC-LNs could be a viable delivery system for site-specific treatment of PAH.