ABSTRACT
Red blood cells (RBCs) are one of the most needed blood products in clinical blood transfusion treatment. At present, RBCs are mainly provided by blood donors, and the contradiction between supply and demand is extremely prominent. Customizable RBCs cultured in vitro bring dawn to solve this contradiction. RBCs can be differentiated and cultured from human induced pluripotent stem cells, immortalized adult erythroid progenitor cells, embryonic stem cells (ESCs), hematopoietic stem cells or the mononuclear cells extracted from umbilical cord blood and peripheral blood. Each method, with its own advantages and drawbacks, are still in the stage of laboratory research. However, according to the current progress, it is not far from clinical application. This paper summarizes the progress in this field.
ABSTRACT
SummaryThe objectives were to develop an effective protocol for transfection of ovine secondary follicles and to assess the effect of attenuating aquaporin 3 (AQP3) using a small interfering RNA (siRNA-AQP3) on antrum formation and follicular growth in vitro. Various combinations of Lipofectamine® volumes (0.5, 0.75 or 1.0 µl), fluorescent oligonucleotide (BLOCK-iT ™) concentrations (3.18, 27.12 or 36.16 nM) and exposure times (12, 14, 16, 18 or 20 h) were tested. The BLOCK-iT™ was replaced by siRNA-AQP3 in the transfection complex. Ovine secondary follicles were isolated and cultured in vitro for 6 days using standard protocols. Follicles were transfected on day 0 or 3 or on both days (0 and 3) and then cultured for an additional 3 or 6 days. As revealed by the fluorescence signal, the Lipofectamine®/BLOCK-iT™ complex (0.75 µl + 27.12 nM by 12 h of incubation) crossed the basement membrane and granulosa cell and reached the oocytes. In general, the rate of intact follicles was higher and the rate of antrum formation was lower in transfected follicles compared with control follicles. In conclusion, ovine secondary follicles can be successfully transfected during in vitro culture, and siRNA-mediated attenuation of AQP3 gene reduced antrum formation of secondary follicles.
Subject(s)
Aquaporin 3/genetics , Ovarian Follicle/physiology , Transfection/methods , Animals , Aquaporin 3/metabolism , Cell Culture Techniques , Female , Gene Knockdown Techniques , Lipids , Ovarian Follicle/growth & development , RNA Interference , SheepABSTRACT
Objective To observe the effects of Gandouling on reactive oxygen species (ROS) level, the expression of nuclear factor erythroid 2-related factor 2 (Nrf2) mRNA and protein of neural stem cells of the mice cultured in high concentration copper. Methods The model of neural stem cells of the mice was cultured in vitro with high concentration copper. The experimental rats were randomly divided into blank control group, model group, and Gandouling low-, medium-, and high-dose groups. Each medication group was given relevant concentration of Gandouling serum for gavage. The MTT was adopted to test proliferation level on neural stem cells; flow cytometer was used to examine the change of ROS level in cells; qPCR was used to measure the expression of Nrf2 mRNA;Western blot was used to measure the change of the level of protein Nrf2 in cells. Results Compared with the blank control group, the proliferation rate of neural stem cells was significantly decreased, ROS levels were significantly increased, and Nrf2 gene and protein expression was significantly decreased (P＜0.01). Compared with the model group, neural stem cells proliferation rate was significantly increased, ROS levels were significantly reduced, and Nrf2 gene and protein expression was significantly increased (P＜0.05, P＜0.01). Conclusion Gandouling can promote the proliferation of neural stem cells in mice by reducing ROS content in high copper-loaded mice and up-regulating Nrf2 expression.
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Objective To investigate the interaction between endometrium and embryo under the co-culture condition during the planting window stage. Methods Twenty patients who cancelled transplant for OHSS in the Reproductive Center of the second affiliated hospital of Zhengzhou university were enrolled.The discarded embryos were used to build the embryos co-culture system (Group A), with a single membrane (group B) and a single embryo(group C)as the control group. Levels of LIF and IGF expression and embryo growth were checked on 3, 4,5,6,7 day post-oocyte retrieval. Results Expressions of LIF mRNA and IGF mRNA in Group A were significantly higher than those in Group B. Embryo growth in the Group A was better than that in Group C. Conclusion Co-culture of endometrium and embryo supports the development of embryo and improves the receptivity of endometrium.
ABSTRACT
All organisms regulate their life activities through the biological clock, which makes a variety of activities regular.For example, many physiological activities such as sleep-wake cycle, temperature, heart rate, blood pressure, endocrine and metabolic activity of the kidney and liver are subject to the regulation of circadian rhythms, that is to say they are all under the control of circadian pacemaker.Physiological activity of cell cultured in vitro also possess rhythms.This paper conducts a brief overview of biological clock of cell cultured in vitro and analyzes the molecular mechanism of the biological clock of the neurons and peripheral tissue cell as well as the existing problems, which provide reference for comprehensive interpretation of the molecular mechanism of biological clock.
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Objective To observe the effects of dehydrocorydaline (DHC) on proliferation and collagen secretion of cardiac fibroblasts (CFs) cultured by high glucose;To provide experimental evidence for clinical application of Rhizoma Corydalis. Methods CFs cultured in vitro with high glucose were made into models. Collagenase and trypsin were used for the combine digestion of CFs from ratsbornin 24 h. 2-4 generation CFs were cultured by high glucose (25 mmol/L), and then 100, 50, 25 mg/L dethydrocorydaline was added for intervention. Cellular morphology of CFs was observed after 24, 48 h. CFs proliferation was detected by MTT method. Cell cycle was assessed via flow cytometry. The levels of collagen Ⅰ and collagen Ⅲ were determined by ELISA. Results CFs began to grow adherence 3 hours after planting, and CFs cultured by high glucose significantly proliferated 24, 48 h later (P<0.05, P<0.01). The percentage of S+G2+M phase CFs increased significantly after 48 h (P<0.01). The secretion of collagen Ⅰ and collagen Ⅲ also increased significantly (P<0.01). After the intervention of DHC, CFs proliferation was significantly inhibited (P<0.01);the percentage of S+G2+M phase CFs decreased (P<0.01);the secretion of collagen Ⅰ and collagen Ⅲ was reduced (P<0.05, P<0.01). Conclusion DHC can reduce CFs proliferation, decrease collagen secretion of CFs cultured by high glucose, and has potential effects of anti-myocardial fibrosis.
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Objective To isolate and identify pancreatic stem cells of Kunming mouse and to observe the differentiation potency of those cells. Methods Pancreas of postnatal Kunming mice were digested by enzymes to isolate pancreatic stem cells. The potency of the cell differentiation was then identified with special marker of cytokeratin 19(CK 19), insulin and glucagons by immunocytochemical staining. Results Few epithelioid cells could be found to survive at the beginning of isolation, but when medium was replaced by that with growth factor, the cells proliferated quickly in fusiform shape and formed colony gradually. The cells were CK 19 immunoreactive positive after transfer of culture. After differentiation induced by high glucose, the cells formed the pancreatic islet like structures. Immunocytochemical staining showed that part of the cells of pancreatic islet like structures were insulin immunoreactive positive and few of the cells were glucagon immunoreactive positive. Conclusion Pancreatic stem cells of Kumming mouse can proliferate when cultured in vitro and have the potency of differentiation into ? and? cells of pancreatic islet.
