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1.
Biotechnol Bioeng ; 121(1): 341-354, 2024 01.
Artigo em Inglês | MEDLINE | ID: mdl-37749931

RESUMO

Recombinant adeno-associated virus (rAAV) is among the most commonly used in vivo gene delivery vehicles and has seen a number of successes in clinical application. Current manufacturing processes of rAAV employ multiple plasmid transfection or rely on virus infection and face challenges in scale-up. A synthetic biology approach was taken to generate stable cell lines with integrated genetic modules, which produced rAAV upon induction albeit at a low productivity. To identify potential factors that restrained the productivity, we systematically characterized virus production kinetics through targeted quantitative proteomics and various physical assays of viral components. We demonstrated that reducing the excessive expression of gene of interest by its conditional expression greatly increased the productivity of these synthetic cell lines. Further enhancement was gained by optimizing induction profiles and alleviating proteasomal degradation of viral capsid protein by the addition of proteasome inhibitors. Altogether, these enhancements brought the productivity close to traditional multiple plasmid transfection. The rAAV produced had comparable full particle contents as those produced by conventional transient plasmid transfection. The present work exemplified the versatility of our synthetic biology-based viral vector production platform and its potential for plasmid- and virus-free rAAV manufacturing.


Assuntos
Células Artificiais , Dependovirus , Dependovirus/genética , Linhagem Celular , Transfecção , Vetores Genéticos
2.
AAPS PharmSciTech ; 23(1): 58, 2022 Jan 20.
Artigo em Inglês | MEDLINE | ID: mdl-35059874

RESUMO

To prevent the sticking of Corni fructus extract (CFE) during spray drying, the anti-sticking effects of different excipients were compared. Hydroxypropyl methylcellulose (HPMC)-VLV showed a higher powder yield at a lower dosage (8% of total solids), and a lower solution viscosity, compared with HPMC-E5. Therefore, HPMC-VLV is a more effective excipient for reducing CFE sticking during spray drying. The spray-drying process parameters were optimized by central composite rotatable design/response surface methodology, and spray drying was conducted under the following conditions: Inlet air temperature, 126 °C; atomization pressure, 1.05 bar; pump speed, 7.7 mL/min. Scanning electron microscopy showed that the powder comprised shrunken spherical particles with particle sizes in the range of 2-30 µm. Analysis of dynamic surface tension and chemical elements on the powder surface showed that HPMC-VLV rapidly moved to the droplet surface owing to its surface activity. HPMC covered the droplet surface and reduced surface tension, achieving an anti-sticking effect. In conclusion, HPMC-VLV at a solid content of 8% significantly improved the spray drying and reduced sticking of CFE. The spray-drying process parameters were nonlinearly related to the dry product yield. Graphical Abstract.


Assuntos
Cornus , Derivados da Hipromelose , Metilcelulose , Extratos Vegetais , Secagem por Atomização
3.
Electrophoresis ; 42(14-15): 1436-1449, 2021 08.
Artigo em Inglês | MEDLINE | ID: mdl-33908064

RESUMO

Profiling of lipid-water partition coefficients (KL/W ) of drugs is an essential issue during the early stage of drug development. In this study, two liposomes, including 1,2-distearoyl-sn-glycero-3-phosphocholine (DSPC) + cholesterol (Chol) (DSPC/Chol liposomes) and soybean lecithin (SPC) + Chol (SPC/Chol liposomes), were prepared for the liposome electrokinetic chromatography (LEKC) analysis, and the logarithm of lipid-water partition coefficients (log KL/W ) of neutral and ionic drugs were determined based on an iterative method. The log KL/W values determined by the SPC/Chol or DSPC/Chol liposomes LEKC were linearly fitted, which showed a good fitting coefficient (R2 = 0.89). Furthermore, the linear relationship between the data obtained from LEKC system and octanol-water system, immobilized artificial membrane, Caco-2 cell model, and software prediction was analyzed, respectively. Results illustrated that DSPC/Chol liposomes or SPC/Chol liposomes had a good linear relationship with Caco-2 cell model, and R2 was 0.81 and 0.72, respectively. Moreover, the linear free energy relationship analysis suggested that the solute volume, hydrogen bond basicity, and J- were the main descriptors that drove the partition process of solutes in the SPC/Chol or DSPC/Chol LEKC system. In addition, the normalized properties of the SPC/Chol and DSPC/Chol LEKC systems through linear free energy relationship analysis were very close. In short, DSPC/Chol liposomes are more suitable for simulating cell membranes than SPC/Chol liposomes, and the developed LEKC is an effective partitioning model for measuring the log KL/W of drugs.


Assuntos
Cromatografia , Células CACO-2 , Colesterol , Humanos , Íons , Cinética , Lipossomos , Membranas Artificiais , Soluções , Água
4.
Drug Metab Dispos ; 47(3): 227-233, 2019 03.
Artigo em Inglês | MEDLINE | ID: mdl-30567880

RESUMO

Anacetrapib is an inhibitor of cholesteryl ester transfer protein (CETP), associated with reduction in LDL cholesterol and increase in HDL cholesterol in hypercholesterolemic patients. Anacetrapib was not taken forward into filing/registration as a new drug for coronary artery diease, despite the observation of a ∼9% reduction in cardiovascular risk in a large phase III cardiovascular outcomes trial (REVEAL). Anacetrapib displayed no adverse effects throughout extensive preclinical safety evaluation, and no major safety signals were observed in clinical trials studying anacetrapib, including REVEAL. However, anacetrapib demonstrated a long terminal half-life in all species, thought to be due, in part, to distribution into adipose tissue. We sought to understand the dependence of anacetrapib's long half-life on adipose tissue and to explore potential mechanisms that might contribute to the phenomenon. In mice, anacetrapib localized primarily to the lipid droplet of adipocytes in white adipose tissue; in vitro, anacetrapib entry into cultured human adipocytes depended on the presence of a mature adipocyte and lipid droplet but did not require active transport. In vivo, the entry of anacetrapib into adipose tissue did not require lipase activity, as the distribution of anacetrapib into adipose was-not affected by systemic lipase inhibition using poloaxamer-407, a systemic lipase inhibitor. The data from these studies support the notion that the entry of anacetrapib into adipose tissue/lipid droplets does not require active transport, nor does it require mobilization or entry of fat into adipose via lipolysis.


Assuntos
Adipócitos/metabolismo , Tecido Adiposo/metabolismo , Proteínas de Transferência de Ésteres de Colesterol/antagonistas & inibidores , Gotículas Lipídicas/metabolismo , Oxazolidinonas/farmacologia , Adipócitos/citologia , Tecido Adiposo/citologia , Animais , Linhagem Celular , HDL-Colesterol/sangue , HDL-Colesterol/metabolismo , LDL-Colesterol/sangue , LDL-Colesterol/metabolismo , Meia-Vida , Humanos , Inibidores de Hidroximetilglutaril-CoA Redutases , Hipercolesterolemia/sangue , Hipercolesterolemia/tratamento farmacológico , Lipase/antagonistas & inibidores , Lipase/metabolismo , Lipólise/efeitos dos fármacos , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Oxazolidinonas/administração & dosagem , Oxazolidinonas/uso terapêutico , Poloxâmero/farmacologia , Distribuição Tecidual/efeitos dos fármacos
5.
Molecules ; 24(15)2019 Jul 24.
Artigo em Inglês | MEDLINE | ID: mdl-31344801

RESUMO

In this work, bamboo fibers are chemically modified with NaOH solution of 1, 4, and 7 wt% concentrations at room temperature, respectively, and subsequently the untreated and treated fibers are prepared with epoxy resin for unidirectional composites by hot pressing molding technique. Tensile and micro-bond tests are conducted on the composite specimens to obtain mechanical properties, such as tensile strength and modulus, elongation at break, and interfacial strength. Besides, scanning electron microscopy (SEM) is employed to perform morphological observations for constituent damages. In addition, the influence of alkali concentration on the thermal performance of epoxy-based composites is examined by using differential scanning calorimetry (DSC) and thermogravimetric (TG) analysis. It is found that composite tensile strength reaches the maximum when the alkali concentration is 4%, increased by 45.24% compared with untreated composites. The composite elongation at break increases on increasing the concentration. Inversely, the composite modulus decreases as the concentration increases. Besides, the results demonstrate that the chemical treatment on the fiber surface could improve interface adhesion, as observed from its topography by SEM. Micro-bond test reveals that there is maximum interfacial shear strength when the alkali concentration is 4%, which increases by 100.30% in comparison with the untreated samples. In case of thermal properties, the DSC analysis indicates that the glass transition temperature is maximized at 4% alkali concentration, which is increased by 12.95%, compared to those from unmodified fibers. In addition, TG results show that the 4% concentration also facilitates thermal stability improvement, indicative of superior interfacial bonding.


Assuntos
Fenômenos Químicos , Compostos de Epóxi/química , Sasa/química , Resinas Epóxi/química , Fenômenos Mecânicos , Propriedades de Superfície , Termogravimetria
6.
Adv Exp Med Biol ; 1077: 451-474, 2018.
Artigo em Inglês | MEDLINE | ID: mdl-30357703

RESUMO

In designing biomaterial for regenerative medicine or tissue engineering, there are a variety of issues to consider including biocompatibility, biochemical reactivity, and cellular interaction etc. Mussel-inspired biomaterials have received much attention because of its appealing features including strong adhesiveness on moist surfaces, enhancement of cell adhesion, immobilization of bioactive molecules and its amenability to post-functionalization via catechol chemistry. In this review chapter, we give a brief introduction on the basic principles of mussel-inspired polydopamine coating, catechol conjugation, and discuss how their features play a vital role in biomedical application. Special emphasis is placed on tissue engineering and regenerative applications. We aspire to give readers of this book a comprehensive insight into mussel-inspired biomaterials that can facilitate them make significant contributions in this promising field.


Assuntos
Materiais Biocompatíveis , Bivalves , Engenharia Tecidual , Animais , Adesão Celular , Humanos , Medicina Regenerativa
7.
Molecules ; 23(7)2018 07 07.
Artigo em Inglês | MEDLINE | ID: mdl-29986521

RESUMO

2-Ethyl-3,6-dimethylpyrazine (EDMP) was an alarm pheromone component isolated from the mandibular gland of the red imported fire ant, Solenopsis invicta Buren. Several pyrazine analogues have been previously found to elicit significant alarm responses in S. invicta workers. This study aimed to separate the commercially available 2-ethyl-5(6)-methylpyrazine (EMP), i.e., a mixture of 2-ethyl-6-methylpyrazine (2E6MP) and 2-ethyl-5-methylpyrazine (2E5MP), and to examine both electroantennogram (EAG) and behavioral responses of S. invicta workers to EMP and the purified isomers. HPLC separations were achieved using a polysaccharide chiral stationary phase (Chiralpak AD-H) column with both mobile phases: Cyclohexane/isopropanol, and hexane/isopropanol. A ratio of 99:1 was selected for the separation of EMP at semipreparative level. The structures of the isomers obtained through the cyclohexane/isopropanol mobile phase were confirmed by detailed analyses of 2D-HSQC- and -HMBC-NMR data. The two isomers showed differential methine C⁻H correlations evidenced by 2D-HMBC-NMR spectra. The two concentrated fractions obtained through hexane/isopropanol mobile phase were subjected to EAG test and behavioral bioassay on S. invicta workers. The two HPLC−purified isomers, 2E6MP and 2E5MP, and their mixture (1:1) at same dose elicited similar EAG and alarm responses, indicating that these two isomers are equally active. The 2D-NMR−spectroscopic characterization, and electrophysiological and alarm activities of 2E6MP and 2E5MP were reported here for the first time.


Assuntos
Formigas/fisiologia , Feromônios/química , Pirazinas/química , Animais , Formigas/química , Antenas de Artrópodes/fisiologia , Comportamento Animal/efeitos dos fármacos , Cromatografia Líquida de Alta Pressão , Isomerismo , Estrutura Molecular
8.
J Vasc Surg ; 65(1): 82-90, 2017 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-27498056

RESUMO

OBJECTIVE: To evaluate efficacy, safety, and midterm patency of endovascular treatment of obstructive popliteal artery (PA) disease. METHODS: A retrospective evaluation of patients with atherosclerotic PA disease who underwent percutaneous transluminal balloon angioplasty and provisional stent, based on both conventional and dynamic angiographies, was conducted from June 2011 to June 2014. Forty-three patients were included in the study, and most patients had limited surgical revascularization options. Demographic characteristics, angiographic findings, interventional data, complications, vessel patency, limb salvage rates, and survival rates were analyzed. RESULTS: The median lesion length was 5 cm with 72.1% having total occlusions. The second popliteal segment (P2) was involved most frequently (60.5%, n = 26). Critical limb ischemia was present in 69.8%. The technical success rate was 92.9% (42/43 limbs), with 29 cases requiring adjunctive nitinol stents after balloon angioplasty (47.6% based on conventional angiography, 21.4% based on dynamic angiography, and 4.8% additional stents based on dynamic angiography). Complications included thromboembolism (2.3%), perforation (2.3%), pseudoaneurysm (2.3%), and myocardial infarction (2.3%). Stent fracture was present in three cases (7.1%) during the mean follow-up period of 18.3 months. The baseline ankle-brachial index significantly improved after the intervention, from 0.49 ± 0.11 to 0.92 ± 0.14 (P < .01). The Rutherford-Becker class decreased from 3.95 ± 0.76 to 1.76 ± 0.95 (P < .01) at 12 months. The 1-year primary, primary-assisted, and secondary patency rates were 75.2% ± 6.8%, 82.4% ± 6.0%, and 89.9% ± 4.8%, respectively. The limb salvage and amputation-free survival rates at 12 months were 91.6% and 87.0%, respectively. CONCLUSIONS: Balloon angioplasty with a provisional stent based on dynamic angiography is a feasible, safe, and effective therapy for patients with obstructive PA disease. Although the occurrence of stent fracture is still inevitable, patients with critical limb ischemia who have limited surgical options may get more benefits from the endovascular treatment of PA obstructive diseases.


Assuntos
Angiografia/métodos , Angioplastia com Balão , Aterosclerose/diagnóstico por imagem , Aterosclerose/terapia , Isquemia/diagnóstico por imagem , Isquemia/terapia , Artéria Poplítea/diagnóstico por imagem , Idoso , Idoso de 80 Anos ou mais , Ligas , Amputação Cirúrgica , Angioplastia com Balão/efeitos adversos , Angioplastia com Balão/instrumentação , Angioplastia com Balão/mortalidade , Índice Tornozelo-Braço , Aterosclerose/mortalidade , Aterosclerose/fisiopatologia , China , Estado Terminal , Feminino , Humanos , Isquemia/mortalidade , Isquemia/fisiopatologia , Estimativa de Kaplan-Meier , Salvamento de Membro , Masculino , Artéria Poplítea/fisiopatologia , Valor Preditivo dos Testes , Modelos de Riscos Proporcionais , Desenho de Prótese , Retratamento , Estudos Retrospectivos , Fatores de Risco , Stents , Taxa de Sobrevida , Fatores de Tempo , Resultado do Tratamento , Grau de Desobstrução Vascular
9.
Blood ; 124(5): 771-9, 2014 Jul 31.
Artigo em Inglês | MEDLINE | ID: mdl-24869939

RESUMO

The Philadelphia chromosomal-negative chronic myeloproliferative neoplasms (MPNs) originate at the level of the hematopoietic stem cell (HSC). The protracted clinical course of the MPNs has limited the use of potentially toxic treatment modalities, which may eliminate the responsible malignant clone. Treatment with low doses of RG7112, an orally available small-molecule inhibitor of p53-MDM2, both alone and combined with pegylated interferon α 2a (Peg-IFNα 2a), significantly decreased MPN colony-forming unit-granulocyte macrophage and burst-forming unit-erythroid numbers and preferentially eliminated the total number of JAKV617F(+) MPN hematopoietic progenitor cells. The effects of RG7112 and Peg-IFNα 2a on MPN progenitor cells were dependent on blocking p53-MDM2 interactions and activating the p53 pathway, thereby increasing MPN CD34(+) cell apoptosis. Treatment of polycythemia vera (PV) and primary myelofibrosis (PMF) CD34(+) cells with low doses of RG7112 and Peg-IFNα 2a before their transplantation into immune-deficient mice decreased the degree of donor-derived chimerism as well as the JAK2V617F allele burden, indicating that these drugs can each alone or in combination deplete MPN HSCs. These results provide a rationale for the use of combinations of low doses of RG7112 and Peg-IFNα 2a for the treatment of PV or PMF patients with the intent of altering their natural history.


Assuntos
Antivirais/farmacologia , Células-Tronco Hematopoéticas/metabolismo , Imidazolinas/farmacologia , Interferon-alfa/farmacologia , Janus Quinase 2 , Mutação de Sentido Incorreto , Policitemia Vera/tratamento farmacológico , Polietilenoglicóis/farmacologia , Mielofibrose Primária/tratamento farmacológico , Proteínas Proto-Oncogênicas c-mdm2/antagonistas & inibidores , Administração Oral , Substituição de Aminoácidos , Animais , Feminino , Transplante de Células-Tronco Hematopoéticas , Células-Tronco Hematopoéticas/patologia , Xenoenxertos , Humanos , Janus Quinase 2/genética , Janus Quinase 2/metabolismo , Masculino , Camundongos , Camundongos Endogâmicos NOD , Camundongos SCID , Policitemia Vera/genética , Policitemia Vera/metabolismo , Policitemia Vera/patologia , Mielofibrose Primária/genética , Mielofibrose Primária/metabolismo , Mielofibrose Primária/patologia , Proteínas Proto-Oncogênicas c-mdm2/genética , Proteínas Proto-Oncogênicas c-mdm2/metabolismo , Proteínas Recombinantes/farmacologia
10.
Am J Pathol ; 184(3): 714-20, 2014 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-24378407

RESUMO

Enterovirus 71 (EV71; family Picornaviridae, species human Enterovirus A) usually causes hand, foot, and mouth disease, which may rarely be complicated by fatal encephalomyelitis. We investigated extra-central nervous system (extra-CNS) tissues capable of supporting EV71 infection and replication, and have correlated tissue infection with expression of putative viral entry receptors, scavenger receptor B2 (SCARB2), and P-selectin glycoprotein ligand-1 (PSGL-1). Formalin-fixed, paraffin-embedded CNS and extra-CNS tissues from seven autopsy cases were examined by IHC and in situ hybridization to evaluate viral antigens and RNA. Viral receptors were identified with IHC. In all seven cases, the CNS showed stereotypical distribution of inflammation and neuronal localization of viral antigens and RNA, confirming the clinical diagnosis of EV71 encephalomyelitis. In six cases in which tonsillar tissues were available, viral antigens and/or RNA were localized to squamous epithelium lining the tonsillar crypts. Tissues from the gastrointestinal tract, pancreas, mesenteric nodes, spleen, and skin were all negative for viral antigens/RNA. Our novel findings strongly suggest that tonsillar crypt squamous epithelium supports active viral replication and represents an important source of viral shedding that facilitates person-to-person transmission by both the fecal-oral or oral-oral routes. It may also be a portal for viral entry. A correlation between viral infection and SCARB2 expression appears to be more significant than for PSGL-1 expression.


Assuntos
Encefalomielite/virologia , Enterovirus Humano A/fisiologia , Proteínas de Membrana Lisossomal/metabolismo , Glicoproteínas de Membrana/metabolismo , Tonsila Palatina/virologia , Receptores Depuradores/metabolismo , Replicação Viral , Adolescente , Adulto , Sistema Nervoso Central/virologia , Criança , Epitélio/virologia , Humanos , Inflamação , Masculino , Boca , RNA Viral/genética , Receptores Virais/metabolismo , Adulto Jovem
11.
Connect Tissue Res ; 56(6): 434-9, 2015 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-25943462

RESUMO

AIM: The aim of this study is to evaluate the capacity of polylactid acid (PLA) fibrous membrane seeded with allogeneic rabbit adipose tissue-derived stem cells (ADSCs) to repair urethral defects in a rabbit model. MATERIALS AND METHODS: Rabbit ADSCs were harvested and phenotypically characterized. Twenty-four New Zealand male rabbits with 5-mm urethral mucosal defects were randomly divided into two groups. They underwent urethroplasty either with PLA fibrous membrane seeded with ADSCs (group A) or blank PLA fibrous membrane (group B). At 4 and 6 weeks after urethroplasty, the urethral grafts were collected and analyzed grossly and histologically. The incidence rate of urethrostenosis was measured. RESULTS: The adipose tissue-derived cells in monolayer culture showed a typical morphology of mesenchymal stem cells (MSCs). They were positive for the MSC marker CD44 but negative for lineage markers CD45 and CD105. Six weeks after surgery, the incidence rate of urethrostenosis in group A was significantly lower than that in group B (p < 0.05). In group A, the ADSC-seeded grafts showed a normal urethral architecture with a thickened muscle layer. In contrast, the newly developed urethra in group B demonstrated a fewer number of urothelial layers and scarce or no smooth muscle cells. CONCLUSION: The PLA scaffold seeded with ADSCs is effective in urethral regeneration in a rabbit model. ADSCs may represent a promising source of seed cells for urethral tissue engineering.


Assuntos
Tecido Adiposo/metabolismo , Membranas Artificiais , Poliésteres/farmacologia , Transplante de Células-Tronco , Células-Tronco/metabolismo , Uretra/cirurgia , Tecido Adiposo/patologia , Aloenxertos , Animais , Masculino , Coelhos , Alicerces Teciduais , Uretra/metabolismo , Uretra/patologia
12.
Blood ; 120(15): 3098-105, 2012 Oct 11.
Artigo em Inglês | MEDLINE | ID: mdl-22872685

RESUMO

Interferon (IFN-α) is effective therapy for polycythemia vera (PV) patients, but it is frequently interrupted because of adverse events. To permit the long-term use of IFN, we propose combining low doses of IFN with Nutlin-3, an antagonist of MDM2, which is also capable of promoting PV CD34(+) cell apoptosis. Combination treatment with subtherapeutic doses of Peg IFN-α 2a and Nutlin-3 inhibited PV CD34(+) cell proliferation by 50% while inhibiting normal CD34(+) cells by 30%. Combination treatment with Nutlin-3 and Peg IFN-α 2a inhibited PV colony formation by 55%-90% while inhibiting normal colony formation by 22%-30%. The combination of these agents also decreased the proportion of JAK2V617F-positive hematopoietic progenitor cells in 6 PV patients studied. Treatment with low doses of Peg IFN-α 2a combined with Nutlin-3 increased phospho-p53 and p21 protein levels in PV CD34(+) cells and increased the degree of apoptosis. These 2 reagents affect the tumor suppressor p53 through different pathways with Peg IFN-α 2a activating p38 MAP kinase and STAT1, leading to increased p53 transcription, whereas Nutlin-3 prevents the degradation of p53. These data suggest that treatment with low doses of both Nutlin-3 combined with Peg IFN-α 2a can target PV hematopoietic progenitor cells, eliminating the numbers of malignant hematopoietic progenitor cells.


Assuntos
Imidazóis/farmacologia , Interferon-alfa/farmacologia , Janus Quinase 2/genética , Mutação/efeitos dos fármacos , Piperazinas/farmacologia , Policitemia Vera/tratamento farmacológico , Polietilenoglicóis/farmacologia , Proteínas Proto-Oncogênicas c-mdm2/antagonistas & inibidores , Proteína Supressora de Tumor p53/antagonistas & inibidores , Apoptose/efeitos dos fármacos , Western Blotting , Quimioterapia Combinada , Células-Tronco Hematopoéticas/efeitos dos fármacos , Células-Tronco Hematopoéticas/patologia , Humanos , Técnicas In Vitro , Mutação/genética , Policitemia Vera/genética , Policitemia Vera/patologia , Reação em Cadeia da Polimerase , Proteínas Recombinantes/farmacologia , Células Tumorais Cultivadas
13.
Water Sci Technol ; 70(10): 1610-6, 2014.
Artigo em Inglês | MEDLINE | ID: mdl-25429448

RESUMO

Polyelectrolyte-single wall carbon nanotube (SCNT) composites are prepared by a solution-based method and used as metal-free cathode catalysts for oxygen reduction reaction (ORR) in air-cathode microbial fuel cells (MFCs). In this study, two types of polyelectrolytes, polydiallyldimethylammonium chloride (PDDA) and poly[bis(2-chloroethyl)ether-alt-1,3-bis[3-(dimethylamino)propyl]urea] (PEPU) are applied to decorate the SCNTs and the resulting catalysts exhibit remarkable catalytic ability toward ORR in MFC applications. The enhanced catalytic ability could be attributed to the positively charged quaternary ammonium sites of polyelectrolytes, which increase the oxygen affinity of SCNTs and reduce activation energy in the oxygen reduction process. It is also found that PEPU-SCNT composite-based MFCs show efficient performance with maximum power density of 270.1 mW m(-2), comparable to MFCs with the benchmark Pt/C catalyst (375.3 mW m(-2)), while PDDA-SCNT composite-based MFCs produce 188.9 mW m(-2). These results indicate that PEPU-SCNT and PDDA-SCNT catalysts are promising candidates as metal-free cathode catalysts for ORR in MFCs and could facilitate MFC scaling up and commercialization.


Assuntos
Fontes de Energia Bioelétrica/microbiologia , Catálise , Eletrodos , Nanotubos de Carbono/química , Oxirredução , Oxigênio/química , Polietilenos/química , Compostos de Amônio Quaternário/química
14.
Biotechnol J ; 19(3): e2400051, 2024 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-38479988

RESUMO

Recombinant adeno-associated virus (rAAV) is widely used as an in vivo delivery vector for gene therapy. It is used in a very large dose, and the large quantities required for broad applications present manufacturing challenges. We have developed a synthetic biology platform of constructing cell lines integrated with essential viral genes which can be induced to produce rAAV without plasmid transfection or virus transduction. Through iterative design-construct-characterization cycles, we have showcased the potential of this synthetic cell production system. Systems characterization of the dynamics of viral transcripts and proteins as well as virus assembly and packaging revealed that the expression level and balance of viral genome and capsid protein are keys to not only the productivity but also the full particle content, an important product quality attribute. Boosting cap gene expression by sequential transfection and integration of multiple copies of the cap gene elevated the rAAV titer to levels on a par with traditional plasmid transfection and virus infection. However, overexpression of the cap gene shifted the balance and kinetics of the genome and capsid. We independently tuned the dynamics of genome amplification and capsid protein synthesis by modulating the induction concentration as well as the time profile, and significantly enhanced full particle content while maintaining a high productivity. This strategy of constructing an inducible stable producer cell line is readily adaptable to rAAV vectors of different serotypes and payloads. It can greatly facilitate scalable production of gene therapy vectors.


Assuntos
Células Artificiais , Dependovirus , Dependovirus/genética , Proteínas do Capsídeo/genética , Proteínas do Capsídeo/metabolismo , Vetores Genéticos/genética , Capsídeo , Linhagem Celular
15.
Adv Healthc Mater ; 13(15): e2400049, 2024 06.
Artigo em Inglês | MEDLINE | ID: mdl-38416676

RESUMO

Wound healing and infection remain significant challenges due to the ineffectiveness against multidrug-resistant (MDR) bacteria and the complex oxidative wound microenvironments. To address these issues, thymoquinone-reinforced injectable and thermosensitive TQ@PEG-PAF-Cur hydrogels with dual functions of microenvironment reshaping and photodynamic therapy are developed. The hydrogel comprises natural compound thymoquinone (TQ) and poly (ethylene glycol)-block-poly (alanine-co-phenyl alanine) copolymers (PEG-PAF) conjugated with natural photosensitizer curcumin (Cur). The incorporation of TQ and Cur reduces the sol-to-gel transition temperature of TQ@PEG-PAF-Cur to 30°C, compared to PEG-PAF hydrogel (37°C), due to the formation of strong hydrogen bonding, matching the wound microenvironment temperature. Under blue light excitation, TQ@PEG-PAF-Cur generates significant amounts of reactive oxygen species such as H2O2, 1O2, and ·OH, exhibiting rapid and efficient bactericidal capacities against methicillin-resistant Staphylococcus aureus and broad spectrum ß-lactamases Escherichia coli via photodynamic therapy (PDT). Additionally, Cur effectively inhibits the expressions of proinflammatory cytokines in skin tissue-forming cells. As a result, the composite hydrogel can rapidly transform into a gel to cover the wound, reshape the wound microenvironment, and accelerate wound healing in vivo. This collaborative antibacterial strategy provides valuable insights to guide the development of multifunctional materials for efficient wound healing.


Assuntos
Curcumina , Farmacorresistência Bacteriana Múltipla , Hidrogéis , Staphylococcus aureus Resistente à Meticilina , Cicatrização , Hidrogéis/química , Hidrogéis/farmacologia , Cicatrização/efeitos dos fármacos , Animais , Curcumina/farmacologia , Curcumina/química , Farmacorresistência Bacteriana Múltipla/efeitos dos fármacos , Staphylococcus aureus Resistente à Meticilina/efeitos dos fármacos , Fotoquimioterapia/métodos , Antibacterianos/farmacologia , Antibacterianos/química , Polietilenoglicóis/química , Polietilenoglicóis/farmacologia , Camundongos , Escherichia coli/efeitos dos fármacos , Fármacos Fotossensibilizantes/farmacologia , Fármacos Fotossensibilizantes/química , Espécies Reativas de Oxigênio/metabolismo , Fototerapia/métodos , Humanos
16.
Biomaterials ; 308: 122540, 2024 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-38537343

RESUMO

Helicobacter pylori (H. pylori) infection is a major cause of gastric diseases. Currently, bismuth-based quadruple therapy is widely adopted for eradicating H. pylori infection. However, this first-line strategy faces several challenges such as drug resistance, intestinal dysbacteriosis, and patients' poor compliance. To overcome these problems, an all-in-one therapeutic platform (CLA-Bi-ZnO2@Lipo) that composed of liposomes loading clarithromycin (CLA), Bi, and ZnO2 hybrid nanoparticles was developed for eradicating multidrug-resistant (MDR) H. pylori. The in vitro and in vivo results showed that CLA-Bi-ZnO2@Lipo could target the infection-induced inflammatory mucosa through liposome mediated nanoparticle-tissue surface charge interaction and quickly respond to the gastric acid environment to release CLA, Bi3+, Zn2+, and H2O2. By oral administration per day, the acid triggered decomposition of CLA-Bi-ZnO2@Lipo could significantly increase intragastric pH to 6 within 30 min; The released CLA, Zn2+, and H2O2 further exerted synergistical anti-bacterial effects in which a ∼2 order higher efficacy in reducing MDR H. pylori burden was achieved in comparison with standard quadruple therapy (p < 0.05); The released Zn2+ and Bi3+ could also alleviate mucosal inflammation. Most importantly, the CLA-Bi-ZnO2@Lipo exhibited superior biosafety and nearly no side effects on intestinal flora. Overall, this study developed a highly integrated and safe anti-MDR H. pylori agent which had great potential to be used as an alternative treatment for MDR H. pylori eradication.


Assuntos
Antibacterianos , Bismuto , Claritromicina , Infecções por Helicobacter , Helicobacter pylori , Lipossomos , Helicobacter pylori/efeitos dos fármacos , Infecções por Helicobacter/tratamento farmacológico , Infecções por Helicobacter/microbiologia , Animais , Bismuto/química , Bismuto/uso terapêutico , Bismuto/farmacologia , Antibacterianos/farmacologia , Antibacterianos/uso terapêutico , Claritromicina/farmacologia , Claritromicina/uso terapêutico , Lipossomos/química , Nanopartículas/química , Óxido de Zinco/química , Óxido de Zinco/farmacologia , Humanos , Camundongos , Peróxido de Hidrogênio/metabolismo , Masculino
17.
Biochem Biophys Res Commun ; 431(2): 169-75, 2013 Feb 08.
Artigo em Inglês | MEDLINE | ID: mdl-23313479

RESUMO

Nutlin3, a non-genotoxic agonist of p53, is currently in phase II clinical trials for cancer treatment. However, its effects on normal tissues and cell types remain largely to be determined. Drugs that can selectively target cancer cells as well as cooperate with the p53 pathway are thus greatly needed. Iron-superoxide dismutase (Fe-SOD) is a potential candidate as it selectively targets cancer cells by eliminating the abnormally high levels of reactive oxygen species (ROS) in cancer cells; it also inhibits cancer cell growth by induction of p27. Here, we show evidence that modulating redox and ROS homeostasis cooperates with Nutlin3 to selectively inhibit cancer cells in vitro and in vivo. Co-treatment of Fe-SOD and Nutlin3 showed synergistic inhibition on cancer cells in vitro, and the induction of p27 appeared to be involved. No effects were observed on normal cells. In addition, such co-treatment further exhibited synergistic inhibition on tumor growth in vivo in a murine B16 xenograft model, while the individual treatments only achieved very limited inhibition. Thus, Fe-SOD cooperated with Nutlin3 to selectively inhibit cancer cells in vitro and in vivo.


Assuntos
Imidazóis/administração & dosagem , Neoplasias/tratamento farmacológico , Piperazinas/administração & dosagem , Superóxido Dismutase/administração & dosagem , Animais , Proliferação de Células/efeitos dos fármacos , Inibidor de Quinase Dependente de Ciclina p27/biossíntese , Sinergismo Farmacológico , Feminino , Células Hep G2 , Humanos , Lipossomos , Células MCF-7 , Melanoma Experimental/tratamento farmacológico , Camundongos , Espécies Reativas de Oxigênio/metabolismo , Neoplasias Cutâneas/tratamento farmacológico , Proteína Supressora de Tumor p53/genética , Proteína Supressora de Tumor p53/metabolismo , Ensaios Antitumorais Modelo de Xenoenxerto
18.
Blood ; 116(20): 4284-7, 2010 Nov 18.
Artigo em Inglês | MEDLINE | ID: mdl-20625010

RESUMO

Polycythemia vera (PV) treatment with interferon α (IFNα) is frequently limited by dose-related toxicity. PV CD34(+) cells are characterized by overexpression of Bcl-xL, which can be antagonized by ABT-737 leading to apoptosis. We explored the effects of ABT-737 and IFNα on PV hematopoiesis. Both IFNα and ABT-737 alone or in combination had a modest effect on normal hematopoiesis but each individually were able to markedly induce PV CD34(+) cell apoptosis and suppress hematopoietic colony formation. The inhibitory activities of these agents in combination were greater against PV hematopoiesis than either agent alone. The exposure of PV CD34(+) cells to low doses of IFNα and ABT-737 in combination resulted in the reduction of the proportion of JAK2V617F(+) colonies similar to that observed with higher doses of IFNα. These data provide the rationale for combination therapy with low doses of IFNα and a BH3 mimetic for patients with PV.


Assuntos
Substituição de Aminoácidos/genética , Compostos de Bifenilo/farmacologia , Células-Tronco Hematopoéticas/efeitos dos fármacos , Interferon-alfa/farmacologia , Janus Quinase 2/genética , Nitrofenóis/farmacologia , Policitemia Vera/enzimologia , Polietilenoglicóis/farmacologia , Sulfonamidas/farmacologia , Proteína bcl-X/antagonistas & inibidores , Antígenos CD34/metabolismo , Compostos de Bifenilo/uso terapêutico , Células da Medula Óssea/efeitos dos fármacos , Células da Medula Óssea/patologia , Ensaio de Unidades Formadoras de Colônias , Sinergismo Farmacológico , Sangue Fetal/citologia , Células-Tronco Hematopoéticas/enzimologia , Humanos , Interferon alfa-2 , Janus Quinase 2/metabolismo , Nitrofenóis/uso terapêutico , Piperazinas/farmacologia , Piperazinas/uso terapêutico , Policitemia Vera/tratamento farmacológico , Policitemia Vera/patologia , Proteínas Recombinantes , Sulfonamidas/uso terapêutico , Proteína bcl-X/metabolismo
19.
J Mater Sci Mater Med ; 23(3): 711-22, 2012 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-22231270

RESUMO

In degenerative disc disease, an injectable hydrogel can fill a degenerate area completely, reduce the risk of implant migration and subsequent loss of height of the intervertebral disc, and minimise surgical defects. Here, we propose a method of preparing an injectable silk fibroin/polyurethane (SF/PU) composite hydrogel by chemical cross-linking under physiological conditions. Mechanical testing was used to determine the mechanical strength of the hydrogel. The impact of hydrogel height on the biomechanical properties was discussed to estimate the working capacity of the hydrogel for further clinical application. Rheological properties were also examined to assess the practical ability of the hydrogel for clinical application. Hydrogel injection and cell assessment is also of interest for clinical application. An SF/PU composite hydrogel can be injected through a small incision. A cell proliferation assay using bone marrow stromal cells showed positive cell viability and increased proliferation over a seven-day period in culture. Importantly, the hydrogel can be monitored in real-time using X-ray fluoroscopy during and after surgery according to the results of X-ray fluoroscopy examination, and shows good visibility based on X-ray assays. In particular, the hydrogel offers the clinically important advantage of visibility in CT and T2-weighted magnetic resonance imaging. Based on the results of the current study, the SF/AU composite hydrogel may offer several advantages for future application in nucleus pulposus replacement.


Assuntos
Fibroínas/química , Hidrogéis , Disco Intervertebral , Poliuretanos/química , Seda/química , Microscopia Eletrônica de Varredura , Reologia
20.
J Pharm Biomed Anal ; 209: 114513, 2022 Feb 05.
Artigo em Inglês | MEDLINE | ID: mdl-34883420

RESUMO

Two liposomes, including 1,2-distearoyl-sn-glycero-3-phosphocholine (DSPC) + 1,2-dimyristoyl-sn-glycero-3-phosphoethanolamine (PE) + cholesterol (Chol) (DSPC/PE/Chol liposomes) and soybean lecithin (SPC) + PE + Chol (SPC/PE/Chol liposomes), were prepared and fixed on the inner wall of capillary column by using the adhesion of polydopamine (PDA) membrane and the cross-linking property of glutaraldehyde (GA). The immobilized liposome capillary column (ILCC) has good repeatability and stability based on the electrophoretic mobility of analyte. A CE method based on the immobilized liposome capillary column chromatography (ILCCC) was successfully developed to study the retention behavior of drugs on ILCC, and the logarithm of retention factor (log k) of neutral and ionic drugs were determined. The results show that the log k measured by the ILCCC based on two liposomes have a good linear fitting (R2 = 0.86). Moreover, the linear relationship between ILCCC system and other related research systems (octanol-water system and immobilized artificial membrane (IAM)) was analyzed, and the results indicate that SPC/PE/Chol ILCCC, DSPC/PE/Chol ILCCC and IAM systems have good fitting results, R2 values are 0.86 and 0.78, respectively. In addition, the normalization coefficients of ILCCC and IAM systems obtained by the linear free energy relationship (LFER) analysis are close and the d value is small. In short, the ILCCC is a simple and feasible method for studying drug membrane permeability.


Assuntos
Lipossomos , Fosfatidiletanolaminas , Permeabilidade da Membrana Celular , Colesterol , Permeabilidade
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