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While hyperthermia has been shown to induce a variety of cytotoxic and sensitizing effects on cancer tissues, the thermal dose–effect relationship is still not well quantified, and it is still unclear how it can be optimally combined with other treatment modalities. Additionally, it is speculated that different methods of applying hyperthermia, such as water bath heating or electromagnetic energy, may have an effect on the resulting biological mechanisms involved in cell death or in sensitizing tumor cells to other oncological treatments. In order to further quantify and characterize hyperthermia treatments on a cellular level, in vitro experiments shifted towards the use of 3D cell spheroids. These are in fact considered a more representative model of the cell environment when compared to 2D cell cultures. In order to perform radiofrequency (RF)-induced heating in vitro, we have recently developed a dedicated electromagnetic field applicator. In this study, using this applicator, we designed and validated an experimental setup which can heat 3D cell spheroids in a conical polypropylene vial, thus providing a reliable instrument for investigating hyperthermia effects at the cellular scale. [ABSTRACT FROM AUTHOR]

To explore the therapeutic effects of aloe polysaccharides (APS) on osteoarthritis (OA) in rats through in vitro and in vivo experiments. Methods The articular chondrocytes of SD suckling mice were isolated in vitro and cultured to 2 generations. The concentration of APS in the experimental group was screened by CCK-8. The chondrocyte proliferation and matrix degradation were assessed by detecting DNA content and proteoglycan (GAG) secretion. The realtime fluorescent quantitative PCR was used to detect osteoarthritis-related gene expression, and enzyme-linked immunosorbent assay (ELISA) method was used to detect inflammatory factor levels. Hematoxylin -Eosin (HE) staining, Safranin O and immunofluorescence staining were used to observe the anti-inflammatory and cartilage protection effects of APS. Its effects on osteoarthritis were further explored in SD rat model. Results The APS concentrations of 5 g/L, 10 g/L and 20 g/L were selected by CCK-8 as the low, medium and high dose experimental groups. It was found that DNA contents were significantly higher in each experimental group than those of the model group (P＜0.05). The detection showed that of GAG secretion was significantly higher in the high-dose experimental group than that of the model group (P＜0.05). PCR results showed that the expression levels of aggrecan (ACAN) and type Ⅱ collagen (COL2A1) were significantly up-regulated in the high-dose experimental group compared with those of the model group (P＜0.05), while tumor necrosis factor (TNF)- α, interleukin (IL)-6, the expression of matrix metalloproteinase (MMP)-3 and MMP-13 were significantly down-regulated (P＜0.05). The secretion levels of TNF-α and IL-6 were significantly lower in the high-dose experimental group than those of the model group (P＜0.05). HE and Safranin O staining also showed that the cell morphology was better in the APS experimental group than that of the model group. The immunofluorescence assay showed that the expression level of MMP13 was significantly lower in the high-dose experimental group than that of the model group (P＜0.05). In vivo articular cartilage tissue Safranin O staining and the International Osteoarthritis Research Society (OARSI) cartilage damage score also showed that the cartilage damage was significantly lower in the experimental group than that in the model group (P＜ 0.05). Conclusion APS has certain anti-inflammatory and cartilage protective effects on OA. [ABSTRACT FROM AUTHOR]

BACKGROUND: A large number of studies on neural stem cell culture have been carried out in aborted fetus, fetal and adult mice, but there are few studies on adult human neural stem cell culture. OBJECTIVE: With the improvement of primary cell culture, to culture and identify adult autologous neural stem cells from the abandoned brain tissues around hematoma cavity during surgery in patients with aneurysm rupture, and then to study the cell cryopreservation and recovery so as to establish the adult autologous neural stem cell bank. METHODS: Over 500 mg abandoned brain tissues were collected during surgery from the surrounding tissues of hematoma cavity in three cases of aneurysm rupture. Adult autologous neural stem cells were cultured in serum-free medium with small tissue piece culture method. Cell growth was investigated by inverted phase contrast microscope. The expression of nestin, the specific antigen of neural stem cells, was confirmed by immunofluorescence method. Culture medium containing 4% fetal bovine serum was used to induce adherent growth of neural stem cells. Subculture was carried out when the cells proliferated to a certain number. Neural stem cells partially cryopreserved were used to establish the adult autologous neural stem cell bank. Recovered cells continued to subculture and induce differentiation. The cell differentiation was induced in the culture medium containing 10% fetal bovine serum. Expression of glial fibrillary acid protein (GFAP; marker for astrocytes), β-tublin (marker for neurons) and SOX10 (marker for oligodendrocytes) was detected by immunofluorescence method. RESULTS AND CONCLUSION: Nestin-positive neurospheres were successfully obtained from the abandoned brain tissues in the three cases through the improved serum-free medium culture. The cells could be amplified in vitro and handed from generation to generation. When cultured in the medium containing 10% fetal bovine serum, neurospheres were differentiated into β-tublin-positive neurons, SOX10-positive oligodendrocytes and GFAP-positive astrocytes. After subculture, passage 5 cells were still positive for nestin and maintained their characteristics of stemness. After recovery, the cryopreserved cells could grow well, still be positive for Nestin and be passed and amplified continuously. It is proved that neural stem cells can be obtained from abandoned brain tissues in different regions (lower frontal lobe and temporal lobe) around the hematoma cavity in patients with aneurysm rupture. These cells can differentiate into neurons, oligodendrocytes and astrocytes. Autologous neural stem cells transplantation is likely to promote the repair of neural function from laboratory to clinical application. [ABSTRACT FROM AUTHOR]

BACKGROUND: The cardiomyocytes obtained by traditional culture methods exhibit poor purity and viability, cells grow in clusters, the connection between cells is less, and most of cells beat separately. OBJECTIVE: To obtain high-purity cardiomyocytes presenting with a sheet-like growth by modified culture method. METHODS: Myocardial tissues were digested using 0.1% trypsin at 4 ℃ overnight, and then digested repeatedly through the addition of type II collagenase, and finally were isolated and purified by differential adhesion method combined with chemical inhibitors. The morphology of cardiomyocytes was observed under inverted phase contrast microscope. The purity of cardiomyocytes was identified by cardic troponin T antibody after cultured for 48 hours. The beating frequency of cardiomyocytes was recorded every two days for 10 consecutive days. RESULTS AND CONCLUSION: The cardiomyocytes adhered within 24 hours of culture, distributed in clusters at 72 hours and grew in sheets at 5-7 days of culture. The survival rate of cardiomyocytes was 96% and the purity identified by immunofluorescence was over 97%. There was no significant difference in the beating frequency at different time. These results suggest that the high-purity and high-viability cardiomyocytes are obtained by the modified culture method, which is an effective method for culturing neonatal cardiomyocytes. [ABSTRACT FROM AUTHOR]

BACKGROUND: Neural stem cell transplantation is an emerging therapeutic option in the recovery of neural lesions and neurodegenerative diseases. Neural stem cell culture and differentiation lay a foundation for the further study. OBJECTIVE: To improve the techniques for the isolation, cultivation, differentiation and identification of neural stem cells, and to explore the biological characteristics of cells. METHODS: The neural stem cells from C57BL/6 fetal rats were isolated and cultured in vitro using neurophere culture method followed by morphological and ultrastucture examination. The growth curve and cell cycle of passage 3 cells were drawn and analyzed. Nestin expression was tested by immunofluorescence. Neural stem cells induced in 1% and 10% fetal bovine serum were identified using anti-GFAP, anti-βIII-tubulin and anti-MBP by immunofluorescence. RESULTS AND CONCLUSION: The neurospheres exhibited strong cell proliferation ability. Under transmission electron microscope, there was a high nuclear/cytoplasmic ratio in the neural stem cells, indicating a low differentiation degree. Immunofluorescence analysis revealed that neural stem cells were positive for Nestin. The induced cells were positive for GFAP, βIII-tubulin, and MBP, indicating these cells were induced to differentiate into astrocytes, neurons and oligodendrocytes, and there were more neurons in 1% fetal bovine serum than those in 10% fetal bovine serum. In conclusion, we could successfully isolate neural stem cells in C57BL/6 mice, and low concentration of fetal bovine serum contributes to more neurons differentiated from neural stem cells. [ABSTRACT FROM AUTHOR]

The article details a study which examined the cryopreservation of A72(dog), CRFK(cat), NB324K(human), MCF7(human), WI38(human), and C636(mosquito) cells using serum free crypreservative. The viability of cells stored in a frozen state while attached to substratum of 25 cm2 culture flasks for 30 to 180 days at -80°C is demonstrated. Study authors claim that their technique provides significant time and cost savings in laboratories that rapidly process numerous cell lines.

The article focuses on a study which explored two aspects of cryopreserving culture mammalian cells during routine laboratory maintenance. The study made use of a serum-free cryopreservative which was proven to be nearly as efficient as the serum-containing preservatives. It also compared cryostage of those cells in suspended versus a pellet form using both techniques.

As the prevalence and incidence of ischemic heart disease continue to increase, so does interest in ischemic heart failure management. Limitations of current therapies have led to research aimed at regenerating and repairing ischemically damaged myocardium through stem-cell therapy. Cell types being evaluated include embryonic stem cells, fetal and neonatal cardiomyocytes, skeletal myoblasts, bone marrow stem cells, peripheral blood CD34+ cells, endothelial progenitor cells, cardiac progenitor cells, and fibroblasts. Preclinical animal studies and promising early results of clinical trials now under way suggest that stem-cell therapy may soon become an important new tool in heart failure management. [ABSTRACT FROM AUTHOR]

The anti-atherogenic properties of 3-hydroxy-3-methylglutaryl coenzyme A (HMG-CoA) reductase inhibitors (statins)have been well established in several circulatory beds. Increasing evidence suggests that statins may help attenuate ischemia-reperfusion injury, a beneficial effect that may be related to the antioxidant capabilities of statins; however, this remains controversial. We performed this study to determine whether the HMG-CoA reductase inhibitor cerivastatin can prevent oxidative stress-induced injury in cultured human aortic endothelial cells (HAEC). The HAEC were subjected to oxidative stress in the absence and presence of increasing concentrations of cerivastatin (50 nM -1,000 nM).Oxidative stress was induced by increasing concentrations of hydrogen peroxide or endogenous superoxide anions generated by the inhibition of superoxide dismutase using diethylthiocarbamate 10 mM).Cell viability and mitochondrial activity were measured by mitochondria-dependent 3-(4,5-dimethyl thiazol-2-yl)-2,5-diphenyl tetrazolium bromide (MTT)conversion. Cell morphology was also examined using light microscopy. Exposing HAEC to cerivastatin for 24 hours had no effect on cell viability using both cell morphology and MTT conversion: the HAEC incubated in 100 nM cerivastatin had 90% ± 2.2% viability of the control. As expected, hydrogen peroxide produced a concentration-dependent decrease in cell viability. Varying concentrations of cerivastatin pretreatment for <18 hours showed no protection of HAEC against hydrogen peroxide-induced injury. As a positive control, the prototype antioxidant N-acetyl-L-cysteine was cytoprotective even with the highest hydrogen peroxide concentration. Neither cerivastatin nor N-acetyl-L-cysteine protected HAEC against diethylthiocarbamate-induced oxidative injury at any concentration. In this study, cerivastatin did not protect cultured HAEC against oxidative stress induced by hydrogen peroxide or diethylthiocarbamate. [ABSTRACT FROM AUTHOR]

During in vitro expansion, adult human articular cartilage-derived cells (HACDC) lose their phenotypic stability and capacity to form cartilage in vivo after 4–6 population doublings (PD). Nevertheless, HACDC can be efficiently expanded for up to 20 PD. Here we show that HACDC can generate cartilage, fibrous tissue, skeletal muscle, bone, and adipocytes depending on the balance between phenotypic stability and environmental cues. When 5 × 106 cells were injected intramuscularly into nude mice, early-passage (EP)-HACDC formed cartilage; late-passage (LP)-HACDC formed mostly fibrous tissue, but a limited number of cells contributed to muscle formation. When 0.5 × 106 cells were injected into regenerating mouse muscle, both EP- and LP-HACDC integrated with host myofibers and expressed muscle genes, but a number of EP-HACDC maintained collagen type II expression. HACDC seeded into Collagraft and implanted subcutaneously into nude mice formed scattered bone islands displaying immunoreactivity for human osteocalcin, and expressing human bone-specific genes. Importantly, neither collagen type II transcript nor cartilage tissue was detected at 8 weeks after implantation. Myogenic, osteogenic, and adipogenic differentiation was induced in vitro using specific culture conditions. These findings provide evidence that in vivo tissue formation by HACDC is specified by a balance between environmental cues and the inherent phenotypic stability. [Copyright &y& Elsevier]