Significant Difference In Cell Growth Research Articles

Characterization of Blow-Spun Polyurethane Scaffolds-Influence of Fiber Alignment and Fiber Diameter on Pericyte Growth.

In this study, fibrous polyurethane (PU) materials with average fiber diameter of 200, 500, and 1000 nm were produced using a solution blow spinning (SBS) process. The effects of the rotation speed of the collector (in the range of 200-25 000 rpm) on the fiber alignment and diameter were investigated. The results showed that fiber alignment was influenced by the rotation speed of the collector, and such alignment was possible when the fiber diameter was within a specific range. Homogeneously oriented fibers were obtained only for a fiber diameter ≥500 nm. Moreover, the changes in fiber orientation and fiber diameter (resulting from changes in the rotation speed of the collector) were more noticeable for materials with an average fiber diameter of 1000 nm in comparison to 500 nm, which suggests that the larger the fiber diameter, the better the controlled architectures that can be obtained. The porosity of the produced scaffolds was about 65-70%, except for materials with a fiber diameter of 1000 nm and aligned fibers, which had a higher porosity (76%). Thus, the scaffold pore size increased with increasing fiber diameter but decreased with increasing fiber alignment. The mechanical properties of fibrous materials strongly depend on the direction of stretching, whereby the fiber orientation influences the mechanical strength only for materials with a fiber diameter of 1000 nm. Furthermore, the fiber diameter and alignment affected the pericyte growth. Significant differences in cell growth were observed after 7 days of cell culture between materials with a fiber diameter of 1000 nm (cell coverage 96-99%) and those with a fiber diameter of 500 nm (cell coverage 70-90%). By appropriately setting the SBS process parameters, scaffolds can be easily adapted to the cell requirements, which is of great importance in producing complex 3D structures for guided tissue regeneration.

CRISPR/Cas9-Mediated Knockout of the HuR Gene in U251 Cell Inhibits Japanese Encephalitis Virus Replication.

Human antigen R (HuR) is an RNA-binding protein that regulates the post-transcriptional reaction of its target mRNAs. HuR is a critical factor in cancer development and has been identified as a potential target in many cancer models. It participates in the viral life cycle by binding to viral RNAs. In prior work, we used CRISPR/Cas9 screening to identify HuR as a prospective host factor facilitating Japanese encephalitis virus (JEV) infection. The HuR gene was successfully knocked out in U251 cell lines using the CRISPR/Cas9 gene-editing system, with no significant difference in cell growth between U251-WT and U251-HuR-KO2 cells. Here, we experimentally demonstrate for the first time that the knockout of the HuR gene inhibits the replication ability of JEV in U251 cell lines. These results play an essential role in regulating the replication level of JEV and providing new insights into virus-host interactions and potential antiviral strategies. It also offers a platform for investigating the function of HuR in the life cycle of flaviviruses.

Tolerance of engineered Rhodosporidium toruloides to sorghum hydrolysates during batch and fed-batch lipid production

BackgroundOleaginous yeasts are a promising candidate for the sustainable conversion of lignocellulosic feedstocks into fuels and chemicals, but their growth on these substrates can be inhibited as a result of upstream pretreatment and enzymatic hydrolysis conditions. Previous studies indicate a high citrate buffer concentration during hydrolysis inhibits downstream cell growth and ethanol fermentation in Saccharomyces cerevisiae. In this study, an engineered Rhodosporidium toruloides strain with enhanced lipid accumulation was grown on sorghum hydrolysate with high and low citrate buffer concentrations.ResultsBoth hydrolysis conditions resulted in similar sugar recovery rates and concentrations. No significant differences in cell growth, sugar utilization rates, or lipid production rates were observed between the two citrate buffer conditions during batch fermentation of R. toruloides. Under fed-batch growth on low-citrate hydrolysate a lipid titer of 16.7 g/L was obtained.ConclusionsCitrate buffer was not found to inhibit growth or lipid production in this engineered R. toruloides strain, nor did reducing the citrate buffer concentration negatively affect sugar yields in the hydrolysate. As this process is scaled-up, $131 per ton of hydrothermally pretreated biomass can be saved by use of the lower citrate buffer concentration during enzymatic hydrolysis.Graphical

Micro texturing and laser irradiation, two stimulus of growth and differentiation to neural like cell on the PMMA polymer

Nerve injuries and neurodegenerative disorders remain serious challenges due to the poor results of nerve cell regeneration. The most promising treatment for such injuries is treatments based on stimulating cells to grow and differentiate stem cells in that area into nerve cells, but there are obstacles in controlling the growth and differentiation into fully functional nerve cells and research is ongoing to overcome these obstacles. In this study, polymethyl methacrylate (PMMA) and its laser modified surfaces were used for cell culture and differentiation into neuron-like cells. In order to show the differentiation of PC12 cells into neuron-like cells, the expression levels of the neuronal markers Map2 and Synapsin-1 were examined. The results showed the growth of PC12 on raw PMMA was increased significantly compared to the control group and micrometric parallel grooves lead to a significant difference in cell growth compared to raw PMMA. Also, the simultaneous irradiation of laser to the cell cultured on PMMA caused more cell adhesion on the surface, faster growth and differentiation of PC12 into neuron-like cells.

Milk fermentation with prebiotic flour of Vasconcellea quercifolia A.St.-Hil.

Non-conventional food plants have bioactive compounds and a high nutritional value. Among these, Vasconcellea quercifolia has nutritional benefits, but it is also easy to cultivate and has a low production cost. In this study, the flour from the unripe fruit of V. quercifolia was evaluated in terms of its potential as a prebiotic for the probiotic bacteria Lactobacillus acidophilus and Bifidobacterium lactis. To do so, fermented milk samples were prepared with 2%, 3%, and 6% of flour and 8.25 log CFU/mL of each microorganism. Samples were analyzed in terms of the number of viable cells of L. acidophilus and B. lactis, as well as pH level, total solids, titratable acidity, and texture in the course of 21days of storage at 4ºC. The obtained microbial viability revealed the in vitro symbiotic effect of flour from V. quercifolia on the probiotic strains of L. acidophilus and B. lactis, which reached 10.20 and 11.19 log CFU/mL, respectively, after 21days of storage, showing a significant difference in cell growth of 1.7 and 2.5 log CFU/mL compared with the control. The pH level decreased from 4.8 to 4.5 after storage time, so it did not alter the conditions for the growth of bacteria. The physical and chemical parameters analyzed did not reveal significant differences (p > 0.05), which indicates product stability. Therefore, flour from the unripe fruit of V. quercifolia has a prebiotic property and can be used as a nutritional supplement for L. acidophilus and B. lactis.

Cultivation, cryopreservation and resuscitation of Theileria annulata transformed cells in serum-free media.

Tropical theileriosis is a protozoan disease caused by Theileria annulata that affects cattle in Northern Africa, the Middle East and Asia where vector ticks of the genus Hyalomma occur. Various measures are applied to control the disease, including vaccination with attenuated T. annulata schizonts. Cultivation of T. annulata schizonts is mainly conducted in media containing Fetal Bovine Serum (FBS), which has some disadvantages such as costs, batch- to-batch variation and ethical concerns. In this study, we conducted three experiments to evaluate the ability of (1) T. annulata strains grown in RPMI with 10% FBS (RPMI-FBS) to adapt and grow in serum-free media (i.e., HL-1, RPMI without FBS supplementation, ISF-1, and M199), (2) a T. annulata strain grown in ISF-1 and subsequently frozen in this medium to grow in ISF-1 again after long-term storage in liquid nitrogen, and (3) a T. annulata strain freshly isolated from infected bovine lymphocytes to growin ISF-1, also after cryopreservation. Cell numbers, schizont index, the viability and generation doubling time were calculated in all experiments. In the first experiment, the Hessiene and Beja cell lines from Tunisia previously cultivated in RPMI-FBS and adapted to serum-free media continued to grow significantly better in RPMI-FBS compared to the serum-freemedia. In the second experiment, a Tunisian cell line (Hessiene) cryopreserved in ISF-1 with 5%[v/v] dimethylsulfoxide (DMSO) grewbetter after thawing in RPMI-FBS compared to ISF-1 with a highly significant difference in cell growth (p < 0.001), whereas the third experiment showed that the Ankara cell line had similar growth characteristics in both RPMI-FBS and ISF-1 before and after thawing, with a shorter generation doubling time in ISF-1 than in RPMI-FBS (p = 0.23). Our findings suggest that freshly isolated cells can be propagated, frozen and thawed in serum-free media such as ISF-1, but once cells are adapted to cultivation in the presence of FBS or resuscitated from frozen storage, propagation in serum-free media may not perform as well as cultivation in RPMI-FBS.

Differentiation of adipose tissue-derived stem cells towards vascular smooth muscle cells on modified poly(L-lactide) foils

The aim of our research was to study the behaviour of adipose tissue-derived stem cells (ADSCs) and vascular smooth muscle cells (VSMCs) on variously modified poly(L-lactide) (PLLA) foils, namely on pristine PLLA, plasma-treated PLLA, PLLA grafted with polyethylene glycol (PEG), PLLA grafted with dextran (Dex), and the tissue culture polystyrene (PS) control. On these materials, the ADSCs were biochemically differentiated towards VSMCs by a medium supplemented with TGFβ1, BMP4 and ascorbic acid (i.e. differentiation medium). ADSCs cultured in a non-differentiation medium were used as a negative control. Mature VSMCs cultured in both types of medium were used as a positive control. The impact of the variously modified PLLA foils and/or differences in the composition of the medium were studied with reference to cell adhesion, growth and differentiation. We observed similar adhesion and growth of ADSCs on all PLLA samples when they were cultured in the non-differentiation medium. The differentiation medium supported the expression of specific early, mid-term and/or late markers of differentiation (i.e. type I collagen, αSMA, calponin, smoothelin, and smooth muscle myosin heavy chain) in ADSCs on all tested samples. Moreover, ADSCs cultured in the differentiation medium revealed significant differences in cell growth among the samples that were similar to the differences observed in the cultures of VSMCs. The round morphology of the VSMCs indicated worse adhesion to pristine PLLA, and this sample was also characterized by the lowest cell proliferation. Culturing VSMCs in the differentiation medium inhibited their metabolic activity and reduced the cell numbers. Both cell types formed the most stable monolayer on plasma-treated PLLA and on the PS control. The behaviour of ADSCs and VSMCs on the tested PLLA foils differed according to the specific cell type and culture conditions. The suitable biocompatibility of both cell types on the tested PLLA foils seems to be favourable for vascular tissue engineering purposes.

PDZK1 induces resistance to apoptosis in esophageal adenocarcinoma cells.

Esophageal cancer is a lethal malignancy with a poor prognosis. The incidence of esophageal adenocarcinoma, which develops from Barrett's esophagus (BE), has recently been increasing. In a previous study, we found that PDZK1 expression is higher in long segment BE compared to that in short-segment BE. However, the function of PDZK1 in the mucosa of BE is unclear. Clarify the role of PDZK1 in BE mucosa using PDZK1 overexpressed cells. Human adenocarcinoma-derived OE33 cells were used as a parental cell line and transfected to generate PDZK1 overexpressed OE33 cells (PC cells) or transfected with empty vector as control cells (NC cells). Cell growth of NC and PC cells in 10% fetal bovine serum was evaluated by cell counting. The effect of PDZK1 on proteasome inhibitor (PSI)-induced apoptosis was qualified by fluorescence microscopy and quantified by flow cytometry. Expression of apoptosis-related proteins was evaluated by western blotting. There were no significant differences in cell growth between NC and PC cells. PSI significantly increased apoptosis in NC cells, but not in PC cells. In response to PSI, increased levels of cleaved-caspase3 and decreased pro-caspase3 levels were found in NC cells, but not in PC cells. In NC cells, PSI significantly decreased Bcl-2 expression without affecting Bax levels. In contrast, high expression of both Bcl-2 and Bax was observed in PC cells. Overexpression of PDZK1 protein induces an apoptosis-resistant phenotype in BE cells, which may be a potential therapeutic target.

Human natural killer cell manufacturing using a closed system process for patients with metastatic solid tumors or hematologic malignancies

Background & Aim As potent effectors of the innate immune system natural killer (NK) cells are currently utilized in clinical trials world-wide to treat a variety of hematologic and solid organ malignancies. The ongoing phase I/II study at the National Institutes of Health (08-H-0186) previously utilized open-system processing requiring 60-80 small GREX flasks and single donor AB plasma in the culture medium. We optimized the manufacturing process to generate large numbers of NK cells using closed-system processing with large 5 liter GREX containers and commercially available pooled AB serum. Methods, Results & Conclusion The manufacturing process entails two stages: Preparation of the feeder EBV-transformed lymphoblastoid cell line (LCL) and NK cell culture. LCL are cultured in closed GREX100-CS vessels with XVIVO15 culture media containing 15mM HEPES, 2mM glutamax and 10% of either commercially available pooled AB serum or single donor AB plasma. With one culture vessel capable of generating 8-10 × 108 LCL feeder cells using 600-800mL of culture medium, 3 vessels sufficiently support a full scale culture. LCL number and viability are similar between groups containing serum vs. plasma. In the second stage, NK cell culture is initiated using 3 GREX500-CS with 40 × 106 NK cells and 400 × 106 irradiated LCL feeder cells in 500mL/vessel. Fresh XVIVO20 containing 15mM HEPES, 2mM glutamax, 500IU/mL IL2 and either 10% AB plasma or pooled AB serum is added on days 4, 8, 12 and 17. On day 14, a portion of the cells are harvested and washed for fresh infusion, and the remaining cells are re-seeded into a new vessel for harvest for second infusion on day 19. Day 14 and day 19 harvest generated 10-14 × 109 (1 vessel) and 50-70 × 109 NK cells (4-5 vessels) for fresh infusion, enough to support the target dose level of 1 × 108 (1st harvest) and 5 × 108 (2nd harvest) NK cells/kg. We found no significant differences in cell growth, viability or TRAIL expression when using AB plasma vs. AB serum. However, TRAIL expression was significantly increased when the concentration of the NK cells was kept below 4 × 106 cells/mL during culture. NK cells also demonstrated tumor cell line killing ability in vitro and had increased CD107. In summary, we have developed a new process that improves safety and consistency for culturing NK cells in large numbers for an ongoing clinical trial at the NIH. This process can be easily adapted by cell therapy manufacturing facilities at most academic institutions.

Growth enhancement of bioethanol-producing microbe Clostridium autoethanogenum by changing culture medium composition

The aim of this study was to optimize the culture medium to enhance the growth of Clostridium autoethanogenum, an acetogenic bacterium that converts synthesis gas to ethanol and acetic acid. In order to optimize the culture medium, four components, yeast extract, trace metal solution, vitamin solution, and Na2S·9H2O, were investigated. Yeast extract was found to have the biggest influence on growth, and a concentration of 5 g/L, 10 times higher than that in basal medium, leading to 2.3 times greater growth. At 10 times the concentration in basal medium, trace metal solution inhibited the growth of the acetogen. A change in the concentration of vitamin solution did not yield any significant difference in cell growth. Lastly, growth with Na2S·9H2O showed a tendency to decrease after an initial cell mass increase for concentrations of 0.5 g/L (basal medium condition) and 0.05 g/L. However, at 5 g/L, C. autoethanogenum grew steadily.

Electrochemically induced tribolayer with molybdenum for hip implants: Tribocorrosion and biocompatibility study

As the number of annual Total Hip Replacement (THR) surgeries continues to increase, the longevity of metal based hip implants is a major concern. Tribochemical reactions cause the Cobalt-Chromium-Molybdenum (CoCrMo) hip implant to release wear debris that interacts with decomposed proteins to form a tribolayer. We conducted an electrochemical investigation in order to understand the role of molybdenum in the stability of the tribolayer under mechanical wear and electrochemical corrosion. Tribolayers made of a bovine calf serum (BCS) and bovine calf serum with sodium molybdates (BCS-Mo) were electrochemically deposited on high carbon CoCrMo discs and subjected to corrosion and tribocorrosion experiments under potentiodynamic conditions in a hip-simulator. Potentiodynamic-polarization curve, Electrochemical Impedance Spectroscopy (EIS) tests and surface characterization techniques were carried out. The results indicate an increased resistance to corrosion under mechanical wear by BCS-Mo coated surface which is more distinct when Mo is added into the electrolyte. In addition, biocompatibility evaluation using MG63 osteosarcoma cells on BCS and BCS-Mo coated samples did not show any statistically significant difference in cell growth compared to uncoated CoCrMo discs. These findings suggest a pre-formed electrochemical tribolayer with sodium molybdates may be a promising pre-implantation treatment of THRs to extend the longevity of implants in vivo.

Abstract 3574: Nicotine prevents chemotherapy-induced peripheral neuropathy in vivo, and fails to stimulate the growth of lung cancer cells or interfere with the effectiveness of chemotherapy in vitro

Abstract Chemotherapy has played a significant and unparalleled role in the treatment and survival of cancer patients. However, this pharmacological approach can lead to long-term symptoms of drug toxicity, including chemotherapy-induced peripheral neuropathy (CIPN), a result of peripheral nerve fiber dysfunction or degeneration. CIPN is characterized by sensory symptoms such as numbness, tingling, cold sensitivity, or allodynia, and an overall decrease in quality of life. Paclitaxel (Taxol), a taxane that is commonly used to treat breast, lung, and ovarian cancers, has been found to cause CIPN in 59 to 78% of patients. There is currently no effective preventative or therapeutic treatment for this side effect, which can present either during or after chemotherapy administration. Without the development of an efficacious treatment, CIPN can be a dose-limiting factor for chemotherapy or delay treatment, thereby influencing survival and quality of life. Our studies have revealed that the prototypical nicotinic acetylcholine receptor agonist, nicotine, is capable of reversing and preventing the development of paclitaxel-induced CIPN in vivo, and does not interfere with the cytotoxic properties of paclitaxel in vitro. Male C57BL/6J mice were infused with 24 mg/kg of nicotine via subcutaneous 7-day osmotic minipumps starting 48 hours prior to paclitaxel treatment, which consisted of 8 mg/kg intraperitoneal injections every other day for a total of four injections. The use of von Frey filament testing revealed that nicotine dose-dependently reverses and prevents paclitaxel-induced mechanical allodynia. In regards to the in vitro studies, MTT and MTS colorimetric assays showed that concentrations of nicotine ranging from 0.1 to 10 μM fail to significantly increase or decrease the viability of A549, H460, Lewis lung carcinoma, or human explant lung cancer cells. Similar results were obtained in ovarian cancer cell lines, where no significant difference in cell growth was observed following nicotine treatment of SKOV-3/DDP and OVCAR-3 cells. Most importantly, the paclitaxel-induced decreases in both H460 cell viability and growth were not significantly attenuated by nicotine. Moreover, 1 μM nicotine did not interfere with paclitaxel-induced sub-G1 DNA content or apoptosis of A549 cells. These findings suggest that nicotinic acetylcholine receptors may be promising drug targets for the prevention and treatment of CIPN. Citation Format: Sarah L. Kyte, Wisam Toma, M I. Damaj, Xianjun Fang, David A. Gewirtz. Nicotine prevents chemotherapy-induced peripheral neuropathy in vivo, and fails to stimulate the growth of lung cancer cells or interfere with the effectiveness of chemotherapy in vitro. [abstract]. In: Proceedings of the 107th Annual Meeting of the American Association for Cancer Research; 2016 Apr 16-20; New Orleans, LA. Philadelphia (PA): AACR; Cancer Res 2016;76(14 Suppl):Abstract nr 3574.

Abstract A03: Lymphocytes lead to circulating tumor cell death from 4D model seeded with non-metastatic lung cancer cell line

Abstract Objective: Mouse lung cancer cell lines 393P and 344SQ are derived from KrasLA1/+ p53R172HΔG mice and have different metastatic potential in immunocompetent mice. 393P cells have local primary tumor growth but do not form metastatic lesions when injected into the subcutaneous space of 129sv mice. However, 344SQ cells form a primary tumor and metastatic lesions when injected into the subcutaneous space. Our aim is to determine if lymphocytes play a role in this differential metastatic lung cancer formation between the two cell lines. Methods: Both 393P and 344SQ cells were grown in the ex vivo 4D lung cancer model. We determined the amount of primary tumor growth, the number of circulating tumor cells (CTCs), and metastatic lesion formation. The CTCs from the 4D model seeded with either cells were injected into the tail veins of immunocompetent 129sv mice (wt) or immunodeficient mice (nu/nu). We calculated the Kaplan Meier survival curve. Next, we added lymphocytes derived from a 129sv mouse spleen to an ex vivo 4D lung cancer model seeded with 393P or 344SQ and isolated the CTCs. We plated the CTCs on a petri dish and compared the survival of these cells. Results: Both 393P and 344SQ cells grew in the 4D model. There were no significant differences in the primary tumor growth and circulating tumor cell formation between the two cell lines. There were no significant differences in the number of metastatic lesion formation between the two cell lines (p=0.5). When CTCs from the 4D model seeded with 344SQ were injected into the tail vein of nu/nu mice or wt mice, there were no differences in the survival rates between the two types of mice (p=0.3). However, nu/nu mice that were injected with CTC from the 4D model seeded with 393P were less likely to survive compared to the wt mice (p=0.002). No metastatic lesions formed in the wt mice injected with 393P CTC for 8 months. The CTC from the 4D model seeded with 344SQ and treated with lymphocytes had no significant difference in cell growth when cultured on a petri dish compared to the control group (p=0.9). However, the CTC from the 4D model seeded with 393P treated with lymphocytes had significantly less live cells compared to the control group (p=0.01). Conclusions: Lymphocytes inhibit metastatic lesion formation in 393P cell lines but not 344SQ cell lines. The ex vivo 4D lung cancer model allows for the identification of the cell type that is responsible for inhibition of metastasis. The model can be used to study the role of different components of a tumor's microenvironment in tumor metastasis. Citation Format: Dhruva K. Mishra, Humberto Juarez, Michael J. Thrall, Min P. Kim. Lymphocytes lead to circulating tumor cell death from 4D model seeded with non-metastatic lung cancer cell line. [abstract]. In: Proceedings of the AACR Special Conference on Tumor Metastasis; 2015 Nov 30-Dec 3; Austin, TX. Philadelphia (PA): AACR; Cancer Res 2016;76(7 Suppl):Abstract nr A03.

Abstract 2866: Defining epigenetic vulnerabilities in synovial sarcoma

Abstract Synovial sarcoma constitutes 10% of all soft-tissue sarcomas and arises most frequently in adolescents and young adults. The defining genetic event of synovial sarcoma and main driver of the disease is the translocation of the SS18 gene on chromosome 18q11 to either the SSX1 and SSX2 genes located on chromosome Xp11. The resulting SS18-SSX fusion oncogene lacks a DNA binding domain, but is thought to exert its function via interaction with transcription factors and chromatin remodelers such as polycomb-group chromatin remodeling factors, and components of the SWI/SNF complex. The fact that the fusion oncogene is the defining event in this disease suggests that, on its own, it is able to generate an epigenetic profile that drives tumorigenesis. In this study we expressed SS18-SSX2 in C2C12 myoblasts to recreate the epigenetic events driving synovial sarcoma. C2C12 myoblasts expressing SS18-SSX1/2 did not show significant differences in cell growth however became completely refractory to differentiation to myotubes. In addition, knockdown of SS18-SSX2 in a mouse synovial sarcoma cell line (derived from a mouse model in which the human SS18-SSX2 fusion oncogene is expressed in the myogenic progenitor compartment) led to strong growth arrest and induction of induction of p16 and p21 cell cycle regulators. This results support the notion that the SS18-SSX oncogene drives tumorigenesis by blocking differentiation programs and by repressing tumor suppressor genes to maintain self-renewal of synovial sarcoma cells. To identify epigenetic vulnerabilities that are specifically generated by the SS18-SSX fusion protein we screened two cell lines using an shRNA library targeting epigenetic modulators: the synovial sarcoma cell line mentioned above and C2C12 myoblasts. Three control shRNAs targeting the SS18-SSX2 oncogene were within the top depletion hits in the synovial sarcoma cell line but were neutral in the C2C12 myoblasts, validating the screen performance. To uncover SS18-SSX specific hits we concentrated in genes within this list that specifically depleted in the synovial sarcoma cell line but not in myoblasts. Genes within this list and to which at least two independent shRNAs scored were subjected to extensive one-by one validation. Our results may uncover epigenetic mechanisms required to maintain self-renewal in synovial sarcoma. Citation Format: Ana Banito, Nilgun Tasdemir, Marc Ladanyi, Scott W. Lowe. Defining epigenetic vulnerabilities in synovial sarcoma. [abstract]. In: Proceedings of the 106th Annual Meeting of the American Association for Cancer Research; 2015 Apr 18-22; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Res 2015;75(15 Suppl):Abstract nr 2866. doi:10.1158/1538-7445.AM2015-2866

Sulfur inhibits the growth of androgen-independent prostate cancer in vivo.

Sulfur is a bright yellow crystalline solid at room temperature. The aim of the present study was to investigate the inhibitory effect of sulfur on prostate cancer (PCa) in vivo. Prostate tumors were developed by injecting 22Rv1 or DU-145 PCa cells into sulfur-treated or untreated nude mice. The weight and volume of the tumors were measured. The cancer cells were separated from the tumors, and analyzed for their growth rate and clonogenicity in culture. The expression of PCa-targeted genes was also assessed using real-time polymerase chain reaction. The rate of growth of 22Rv1 tumors in sulfur-treated nude mice gradually decreased, and was reduced by 41.99% (P<0.01) after 22 days when compared with that of the control group. In addition, the growth of DU-145 tumors was also suppressed by 75.16% (P<0.05) after 11 weeks. The clonogenicity of the sulfur-treated tumor cells decreased by 36.7% when compared with that of the control cells. However, no significant difference in cell growth was identified. mRNA levels of the androgen-receptor, prostate specific antigen and human Hox (NKX3.1) genes were significantly decreased by 32.8, 48.2 and 42.2% in sulfur-treated tumors, respectively. Additionally, it was found that the hydrogen sulfide concentration in the serum of sulfur-treated mice was increased by 4.73% (P<0.05). Sulfur significantly suppressed the growth of PCa in vivo. Since sulfur is a known ingredient used in traditional Chinese medicine, it may be used clinically for the treatment of PCa, independently or in combination with other medicine.

Five-year incidence of Nd:YAG laser capsulotomy and association with in vitro proliferation of lens epithelial cells from individual specimens: a case control study.

BackgroundThe aims of this study were to determine the 5-year incidence of posterior capsule opacification (PCO) requiring Nd:YAG laser capsulotomy in a representative mixed cohort of cataract patients, to determine risk factors for PCO and to investigate possible association with growth of human lens epithelial cells (HLEC) in vitro.MethodsPieces of the anterior lens capsule and adhering HLEC were obtained at cataract surgery and cultured individually. After one and two weeks respectively, cultured cells were stained with carboxy-fluorescein diacetate succinimidyl ester (CFDA SE), after which image processing software was used to determine the area of the confluent cell layer. The 5-year incidence of Nd:YAG laser capsulotomy in this cohort was determined through medical records and by mail or telephone interviews. For statistic analyses Mann–Whitney U-test, Fisher’s exact test and binary logistic regression were used.ResultsData on treatment/no treatment for PCO was obtained from 270 patients with a median follow-up time of 57 months (range 50–64 months). The three-year cumulative incidence of PCO was 5.2% and the cumulative 5-year incidence was 11.9%. Patients who had undergone Nd:YAG laser capsulotomy were significantly younger (median 71 years) than patients who did not receive treatment for PCO (median 75 years, p = 0.022). Logistic regression demonstrated that apart from younger age, follow-up time and type of intraocular lens (IOL) were associated with risk of PCO, with hydrophilic 1-piece IOLs conferring a higher risk than hydrophobic acrylic 1-piece or 3-piece IOLs (adjusted OR = 9.4, 95% CI 2.5-35.7, p = 0.001). Of the 270 patients from whom information could be retrieved regarding PCO treatment, in vitro cell culture could be established and quantified from 185 patients. No significant difference in cell growth in vitro was shown between patients subsequently requiring/not requiring Nd:YAG laser capsulotomy.ConclusionsThe cumulative 5-year incidence of 11.9% is comparable or slightly higher than reported in other recent studies. The type of IOL was the most important risk factor for PCO in this study, whereas intrinsic proliferative capacity of the individual’s lens epithelial cells seems to be less important for subsequent PCO development.

A xenogeneic‐free bioreactor system for the clinical‐scale expansion of human mesenchymal stem/stromal cells

The large cell doses (>1 × 10(6) cells/kg) used in clinical trials with mesenchymal stem/stromal cells (MSC) will require an efficient production process. Moreover, monitoring and control of MSC ex-vivo expansion is critical to provide a safe and reliable cell product. Bioprocess engineering approaches, such as bioreactor technology, offer the adequate tools to develop and optimize a cost-effective culture system for the rapid expansion of human MSC for cellular therapy. Herein, a xenogeneic (xeno)-free microcarrier-based culture system was successfully established for bone marrow (BM) MSC and adipose tissue-derived stem/stromal cell (ASC) cultivation using a 1L-scale controlled stirred-tank bioreactor, allowing the production of (1.1 ± 0.1) × 10(8) and (4.5 ± 0.2) × 10(7) cells for BM MSC and ASC, respectively, after 7 days. Additionally, the effect of different percent air saturation values (%Airsat ) and feeding regime on the proliferation and metabolism of BM MSC was evaluated. No significant differences in cell growth and metabolic patterns were observed under 20% and 9%Airsat . Also, the three different feeding regimes studied-(i) 25% daily medium renewal, (ii) 25% medium renewal every 2 days, and (iii) fed-batch addition of concentrated nutrients and growth factors every 2 days-yielded similar cell numbers, and only slight metabolic differences were observed. Moreover, the immunophenotype (positive for CD73, CD90 and CD105 and negative for CD31, CD80 and HLA-DR) and multilineage differentiative potential of expanded cells were not affected upon bioreactor culture. These results demonstrated the feasibility of expanding human MSC from different sources in a clinically relevant expansion configuration in a controlled microcarrier-based stirred culture system under xeno-free conditions. The further optimization of this bioreactor culture system will represent a crucial step towards an efficient GMP-compliant clinical-scale MSC production system.

The Influence of Nuclear Magnetic Resonance Therapy (NMRT) and Interleukin IL1-β Stimulation on Cal 78 Chondrosarcoma Cells and C28/I2 Chondrocytes

The Influence of Nuclear Magnetic Resonance Therapy (NMRT) and Interleukin IL1-β Stimulation on Cal 78 Chondrosarcoma Cells and C28/I2 Chondrocytes

Cyclic strain amplitude dictates the growth response of vascular smooth muscle cells in vitro: role in in-stent restenosis and inhibition with a sirolimus drug-eluting stent

The putative effects of changes in mean strain and cyclic strain amplitude on vascular smooth muscle cell (vSMC) growth (proliferation and apoptosis) were examined. Subsequently, a quantitative measure of vSMC growth was obtained to determine the prolonged effect of changes in mechanical burden following bare-metal stent (BMS) and sirolimus drug-eluting stent (DES) deployment in vitro. Bovine aortic vSMCs were exposed to prolonged cyclic strain using a Flexercell(TM) Tension system and a novel Sylgard(TM) phantom vessel following stent implantation before the level of vSMC proliferation and apoptosis was assessed by FACS analysis, cell counting, and immunocytochemistry. Physiological cyclic strain (5%) decreased vSMC proliferation and increased apoptosis in a temporal manner. There was no significant difference in cell growth following exposure to varying mean strains with similar amplitude. In contrast, exposure to varying strain amplitudes with similar mean strains resulted in significant differences in cell proliferation and apoptosis. In parallel studies, the level of vSMC proliferation and cell survival was significantly increased within low amplitude, high mean strain regions of a phantom vessel following BMS implantation when compared to regions of higher strain amplitude upstream and downstream of the stent, respectively. Moreover, the level of vSMC growth within the stented region was significantly attenuated following implantation of a sirolimus-coated DES independent of significant changes in cell survival. Cyclic strain amplitude is an important regulator of vSMC growth capacity within a stent and is a target for inhibition using a sirolimus-coated DES.

Association of Differentiation-Related Gene-1 (DRG1) with Breast Cancer Survival and in Vitro Impact of DRG1 Suppression

Differentiation-related gene-1, DRG1, is a metastasis suppressor gene whose expression has been shown to be dysregulated in a number of malignancies. The current study examines the expression of DRG1 in a clinical breast cohort and its association with a number of clinical pathological factors using quantitative polymerase chain reaction. Additionally, DRG1 expression is targeted in vitro using ribozyme transgene technology to explore the function of DRG1 in two human breast cancer cell lines. Low levels of DRG1 were found in patients who developed metastasis (p = 0.036) and who died of breast cancer (p = 0.0048) compared to disease free patients. Knockdown of DRG1 also resulted in significantly increased invasion and motility, but decreased matrix-adhesion in MCF7 cells. Knockdown of DRG1 seemed to have minimal impact on the cellular functions of the MDA-MB-231 breast cancer cell line causing no significant differences in cell growth, invasion, motility or matrix-adhesion. Thus, DRG1 appears to be linked to development of metastasis and death in patients who died as a result of breast cancer and may be useful as a prognostic factor as its knockdown appears to be linked with increased invasion and motility and decreased adhesion in MCF7 breast cancer cells.