Gene therapy strategies and prospects for neurofibromatosis type 1 / 中国修复重建外科杂志

OBJECTIVE@#To summarize the gene therapy strategies for neurofibromatosis type 1 (NF1) and related research progress.@*METHODS@#The recent literature on gene therapy for NF1 at home and abroad was reviewed. The structure and function of the NF1 gene and its mutations were analyzed, and the current status as well as future prospects of the transgenic therapy and gene editing strategies were summarized.@*RESULTS@#NF1 is an autosomal dominantly inherited tumor predisposition syndrome caused by mutations in the NF1 tumor suppressor gene, which impair the function of the neurofibromin and lead to the disease. It has complex clinical manifestations and is not yet curable. Gene therapy strategies for NF1 are still in the research and development stage. Existing studies on the transgenic therapy for NF1 have mainly focused on the construction and expression of the GTPase-activating protein-related domain in cells that lack of functional neurofibromin, confirming the feasibility of the transgenic therapy for NF1. Future research may focus on split adeno-associated virus (AAV) gene delivery, oversized AAV gene delivery, and the development of new vectors for targeted delivery of full-length NF1 cDNA. In addition, the gene editing tools of the new generation have great potential to treat monogenic genetic diseases such as NF1, but need to be further validated in terms of efficiency and safety.@*CONCLUSION@#Gene therapy, including both the transgenic therapy and gene editing, is expected to become an important new therapeutic approach for NF1 patients.

Transmembrane mechanism and intracellular trafficking of folic acid-polyethyleneimine carbon dots / 吉林大学学报(医学版)

Objective:To investigate the transport pathway and intracellular distribution of the of fluorescent carbon dots(CDs)synthesized by folic acid and polyethyleneimine(PEI)through the membrane of MC3T3-E1 cells and its effect on the cells,and to clarify the mechanism.Methods:The fluorescent CDs with the function of cell imaging were synthesized by hydrothermal method using folic acid and PEI as the raw materials;MTT assay was applied to screen the best concentration of CDs.The MC3T3-E1 cells were divided into blank control group,folic acid group and CDs group.The biocompatibility of CDs was evaluated by the detection of cell cycle,apoptosis and cellular reactive oxygen species(ROS)level.Nystatin as a kind of caveolae inhibitor and nocodazole as a kind of macropinocytosis inhibitor were used to find out the pathway through which the cells took in the CDs.Using the charcteristic of CDs with blue fluorescence stimulated by ultraviolet ray,the organelle probes were used to observe the distribution of CDs.Results:Compared with blank control group,the cells in different concentrations(100-450 mg·L-1)of CDs groups showed no cytotoxicity at 24 h(P>0.05);at 48 h,the cell proliferation rate was reduced to 68.4% of blank control group when the concentration of CDs reached 350 mg · L -1(P<0.05). Compared with blank control group,the percentages of cells in G0phase and G1phase in CDs group were decreased (P<0.05),and the percentage of cells in S phase was increased(P<0.05);the percentages of cells in G2phase and M phase were increased,but there no was significant differences(P>0.05).Compared with blank control group,the apoptotic rates of the cells in folic acid group and CDs group had no significant differences(P>0.05). Compared with blank control group,the intracellular ROS levels in folic acid group and CDs group were significantly decreased(P<0.05).Compared with blank control group,the uptake amount of CDs in the cells was decreased in nystatin group(P<0.05).The blue fluorescence of CDs overlapped with the red fluorescence of mitochondria under an inverted fluorescence microscope,the blue fluorescence of CDs overlapped with the red fluorescence of lysosomes;they didn't overlap completely with the red fluorescence of the endoplasmic reticulum;the blue fluorescence of CDs overlapped poorly with the red fluorescence of Golgi apparatus.Conclusion:CDs perform well in biocompatibility and they can be distributed to different organelles after taken in by the cells.They can be used as a kind of gene carrier in transgenic therapy.

Transmembrane mechanism and intracellular trafficking of folic acid-polyethyleneimine carbon dots / 吉林大学学报(医学版)

Objective: To investigate the transport pathway and intracellular distribution of the of fluorescent carbon dots (CDs) synthesized by folic acid and polyethyleneimine (PEI) through the membrane of MC3T3-E1 cells and its effect on the cells, and to clarify the mechanism. Methods: The fluorescent CDs with the function of cell imaging were synthesized by hydrothermal method using folic acid and PEI as the raw materials; MTT assay was applied to screen the best concentration of CDs. The MC3T3-E1 cells were divided into blank control group, folic acid grodp and CDs grodp. The biocompatibility of CDs was evaldated by the detection of cell cycle, apoptosis and celldlar reactive oxygen species (ROS) level. Nystatin as a kind of caveolae inhibitor and nocodazole as a kind of macropinocytosis inhibitor were dsed to find od the pathway throdgh which the cells took in the CDs. Using the charcteristic of CDs with bld fldorescence stimdlated by dltraviolet ray, the organelle probes were dsed to observe the distribdtion of CDs. Results: Compared with blank control grodp, the cells in different concentrations (100 - 450 mg · L-1) of CDs grodps showed no cytotoxicity at 24 h (P>0.05); at 48 h, the cell proliferation rate was reddced to 68.4% of blank control grodp when the concentration of CDs reached 350 mg · L-1(P0.05). Compared with blank control grodp, the apoptotic rates of the cells in folic acid grodp and CDs grodp had no significant differences (P>0.05). Compared with blank control grodp, the intracelldlar ROS levels in folic acid grodp and CDs grodp were significantly decreased (P<0.05). Compared with blank control grodp, the dptake amodnt of CDs in the cells was decreased in nystatin grodp (P<0.05). The blde fldorescence of CDs overlapped with the red fldorescence of mitochondria dnder an inverted fldorescence microscope, the blde fldorescence of CDs overlapped with the red fldorescence of lysosomes; they didn't overlap completely with the red fldorescence of the endoplasmic reticdldm; the blde fldorescence of CDs overlapped poorly with the red fldorescence of Golgi apparatds. Conclusion: CDs perform well in biocompatibility and they can be distribdted to different organelles after taken in by the cells. They can be dsed as a kind of gene carrier in transgenic therapy.

Enfermedad de vnn gierke: nuevas tendencias en el manejo / Von gieke diseasse: new trends in management / Doença de vn gieerke: novas tendências no tratamento

La enfermedad de von Gierke, también conocida como enfermedad de deposito de glucógeno tipo Ia, es una enfermedad producida por la deficiencia de la unidad catalítica de la G6Pasa-a, encargada de hidrolizar la glucosa 6 fosfato en el citoplasma celular durante la gluconeogénesis y la glucogenolisis. Las complicaciones a largo plazo son hipoglicemia severa y alteraciones en el crecimiento. En los niños más pequeños la enfermedad típicamente se presenta con crisis convulsivas y hepatomegalia que se manifiestan a los 6 y 8 meses. Otras complicaciones son osteoporosis, gota, enfermedad renal, hipertensión pulmonar y adenomas hepáticos que pueden malignizarse. No se ha encontrado una cura y de no recibir un manejo adecuado es letal en las primeras dos décadas de la vida. El tratamiento consiste en terapia nutricional, asociada a varios medicamentos convencionales. Algunos pacientes pueden requerir transplante renal o transplante hepático. Una nueva esperanza se ha abierto con el advenimiento de la terapia génica con vectores virales, esta estrategia hasta ahora esta siendo desarrollada, pero los estudios realizados han mostrado una luz de esperanza para investigadores, médicos y pacientes. Faltan estudios para que estos tratamientos permitan un beneficio a largo plazo y su aplicación en humanos, ya que las pruebas como es de esperarse solo han sido desarrolladas en modelos animales.

Von Gierke disease, also known as glycogen storage disease type Ia, is a disease caused by deficiency of the G6Pase-a catalytic unit, which hydrolyzes glucose-6- phosphate in the cell cytoplasm during gluconeogenesis and glycogenolysis. Long term complications include severe hypoglycemia and growth disturbances. In small children, the disease typically presents with seizure crisis and hepatomegaly which become manifest at the age of 6 and 8 months. Other complications include osteoporosis, gout, renal disease, pulmonary hypertension and hepatic adenomas which can become malignant. No cure has been found for this disease and it can turn out to be lethal if no appropriate management is given during the first two decades of life. The treatment consists of nutritional therapy associated with a number of conventional drugs. Some patients may require renal or liver transplant. A new hope has emerged with the arrival of gene therapy with viral vectors, strategy that is being developed hitherto, yet performed studies have shown a glimmer of hope for investigators, doctors and patients. There is a need for studies so these treatments allow for a longer term benefit and their application in humans since, as expected, the tests have been developed only in animal models.

A doença de Von Gierke, também conhecida como Glicogenose tipo I, é uma doença produzida pela deficiência da unidade catalítica da G6Pasa-a, encarregada de hidrolisar a glicose 6 fosfato no citoplasma celular durante a gliconeogênese e a glicogenólise. As complicações a longo prazo são hipoglicemia severa e alterações no crescimento. Nas crianças menores a doença se apresenta tipicamente com crises convulsivas e hepatomegalia que se manifestam aos 6 e 8 meses. Outras complicações são osteoporose, gota, doença renal, hipertensão pulmonar e adenomas hepáticos que podem malignizar-se. Não foi encontrada uma cura e se não recebe tratamento adequado é letal nas primeiras duas décadas de vida. O tratamento consiste em terapia nutricional, associada a vários medicamentos convencionais. Alguns pacientes podem requerer transplante renal ou transplante hepático. Uma nova esperança apareceu com a terapia gênica com vetores virais, esta estratégia até agora esta sendo desenvolvida, mas os estudos realizados mostram uma luz de esperança para pesquisadores, médicos e pacientes. Faltam estudos para que estes tratamentos permitam um beneficio a longo prazo e a sua aplicação em humanos, já que os testes como é de se esperar só foram desenvolvidos em modelos animais.