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Effect of culture media on expansion properties of human umbilical cord matrix-derived mesenchymal cells

Salehinejad P, Alitheen NB, Nematollahi-Mahani SN, Ali AM, Omar AR, Janzamin E, et al.

Cytotherapy, 2012 Sep;14(8):948-53.
PMID: 22587592 DOI: 10.3109/14653249.2012.684377

BACKGROUND AIMS: Mesenchymal stromal cells (MSC) have been isolated from a number of different tissues, including umbilical cord. Because of the lack of a uniform approach to human umbilical cord matrix-derived mesenchymal (hUCM) cell expansion, we attempted to identify the optimum conditions for the production of a high quantity of hUCM cells by comparing two media.
METHODS: We compared the ability of Dulbecco's Modified Eagle's Medium/F12 (DMEM/F12) and Alpha Minimum Essential Medium (α-MEM) with Glutamax (GL) (α-MEM/GL) to expand hUCM cells. For this purpose, hUCM cells were cultured in plates containing different culture media supplemented with 10% fetal bovine serum (FBS). Culture dishes were left undisturbed for 10-14 days to allow propagation of the newly formed hUCM cells. The expansion properties, CD marker expression, differentiation potential, population doubling time (PDT) and cell activity were compared between the two groups.
RESULTS: The hUCM cells harvested from each group were positive for MSC markers, including CD44, CD90 and CD105, while they were negative for the hematopoietic cell surface marker CD34. Differentiation into adipogenic and osteogenic lineages was confirmed for both treatments. Cell activity was higher in the α-MEM/GL group than the DMEM/F12 group. PDT was calculated to be 60 h for the DMEM/F12 group, while for the α-MEM/GL group it was 47 h.
CONCLUSIONS: Our data reveal that α-MEM/GL with 10% FBS supports hUCM cell growth more strongly than DMEM/F12 with 10% FBS.

Matched MeSH terms: Cell Culture Techniques*
Cryopreserved mesenchymal stromal cell treatment is safe and feasible for severe dilated ischemic cardiomyopathy

Chin SP, Poey AC, Wong CY, Chang SK, Teh W, Mohr TJ, et al.

Cytotherapy, 2010;12(1):31-7.
PMID: 19878080 DOI: 10.3109/14653240903313966

Bone marrow (BM) mesenchymal stromal cells (MSC) represent a novel therapy for severe heart failure with extensive myocardial scarring, especially when performed concurrently with conventional revascularization. However, stem cells are difficult to transport in culture media without risk of contamination, infection and reduced viability. We tested the feasibility and safety of off-site MSC culture and expansion with freeze-controlled cryopreservation and subsequent rapid thawing of cells immediately prior to implantation to treat severe dilated ischemic cardiomyopathy.

Matched MeSH terms: Cell Culture Techniques
Generating neuron-like cells from BM-derived mesenchymal stromal cells in vitro

Choong PF, Mok PL, Cheong SK, Leong CF, Then KY

Cytotherapy, 2007;9(2):170-83.
PMID: 17453969

The multipotency of stromal cells has been studied extensively. It has been reported that mesenchymal stromal cells (MSC) are capable of differentiating into cells of multilineage. Different methods and reagents have been used to induce the differentiation of MSC. We investigated the efficacy of different growth factors in inducing MSC differentiation into neurons.

Matched MeSH terms: Cell Culture Techniques/methods
Safety and efficacy of dermal fibroblast conditioned medium (DFCM) fortified collagen hydrogel as acellular 3D skin patch

Maarof M, Mh Busra MF, Lokanathan Y, Bt Hj Idrus R, Rajab NF, Chowdhury SR

Drug Deliv Transl Res, 2019 02;9(1):144-161.
PMID: 30547385 DOI: 10.1007/s13346-018-00612-z

Skin substitutes are one of the main treatments for skin loss, and a skin substitute that is readily available would be the best treatment option. However, most cell-based skin substitutes require long production times, and therefore, patients endure long waiting times. The proteins secreted from the cells and tissues play vital roles in promoting wound healing. Thus, we aimed to develop an acellular three-dimensional (3D) skin patch with dermal fibroblast conditioned medium (DFCM) and collagen hydrogel for immediate treatment of skin loss. Fibroblasts from human skin samples were cultured using serum-free keratinocyte-specific media (KM1 or KM2) and serum-free fibroblast-specific medium (FM) to obtain DFCM-KM1, DFCM-KM2, and DFCM-FM, respectively. The acellular 3D skin patch was soft, semi-solid, and translucent. Collagen mixed with DFCM-KM1 and DFCM-KM2 showed higher protein release compared to collagen plus DFCM-FM. In vitro and in vivo testing revealed that DFCM and collagen hydrogel did not induce an immune response. The implantation of the 3D skin patch with or without DFCM on the dorsum of BALB/c mice demonstrated a significantly faster healing rate compared to the no-treatment group 7 days after implantation, and all groups had complete re-epithelialization at day 17. Histological analysis confirmed the structure and integrity of the regenerated skin, with positive expression of cytokeratin 14 and type I collagen in the epidermal and dermal layer, respectively. These findings highlight the possibility of using fibroblast secretory factors together with collagen hydrogel in an acellular 3D skin patch that can be used allogeneically for immediate treatment of full-thickness skin loss.

Matched MeSH terms: Cell Culture Techniques
Effect of extracts from Phyllanthus watsonii Airy Shaw on cell apoptosis in cultured human breast cancer MCF-7 cells

Ramasamy S, Abdul Wahab N, Zainal Abidin N, Manickam S

Exp. Toxicol. Pathol., 2013 Mar;65(3):341-9.
PMID: 22217449 DOI: 10.1016/j.etp.2011.11.005

Species of Phyllanthus have traditionally been used for hundreds of years for treating many ailments including diabetes, anemia, bronchitis and hepatitis. The present study aims to investigate the cytotoxic and apoptotic effects of methanol (PWM), hexane (PWH) and ethyl acetate (PWE) extracts from the leaves of the endemic plant Phyllanthus watsonii Airy Shaw (Phyllanthaceae) on MCF-7 human breast cancer cells. We observed that the PWM, PWH and PWE extracts were cytotoxic and selectively inhibited the growth and proliferation of MCF-7 cells compared to untreated control in a dose dependent manner with an IC(50) of 12.7 ± 4.65, 7.9 ± 0.60 and 7.7 ± 0.29 μg/ml, respectively. However, the extracts were not toxic at these concentrations to normal human lung fibroblast MRC-5 cells. Cell death induced by PWM, PWH and PWE extracts were mainly due to apoptosis which was characterized by apoptotic morphological changes and a nuclear DNA fragmentation. Caspase-3 activation following P. watsonii extracts treatment was also evident for apoptotic cell death which was preceded by an S phase cell cycle perturbation. The results suggested that the cytotoxic activity of P. watsonii extracts was related to an early event of cell cycle perturbation and a later event of apoptosis. Hence, P. watsonii displays potential to be further exploited in the discovery and development of new anticancer agents.

Matched MeSH terms: Cell Culture Techniques
Fulltext Extrinsic factors involved in the differentiation of stem cells into insulin-producing cells: an overview

Wong RS

Exp Diabetes Res, 2011;2011:406182.
PMID: 21747828 DOI: 10.1155/2011/406182

Diabetes mellitus is a chronic disease with many debilitating complications. Treatment of diabetes mellitus mainly revolves around conventional oral hypoglycaemic agents and insulin replacement therapy. Recently, scientists have turned their attention to the generation of insulin-producing cells (IPCs) from stem cells of various sources. To date, many types of stem cells of human and animal origins have been successfully turned into IPCs in vitro and have been shown to exert glucose-lowering effect in vivo. However, scientists are still faced with the challenge of producing a sufficient number of IPCs that can in turn produce sufficient insulin for clinical use. A careful choice of stem cells, methods, and extrinsic factors for induction may all be contributing factors to successful production of functional beta-islet like IPCs. It is also important that the mechanism of differentiation and mechanism by which IPCs correct hyperglycaemia are carefully studied before they are used in human subjects.

Matched MeSH terms: Cell Culture Techniques/methods*
Effects of ranibizumab on the extracellular matrix production by human Tenon's fibroblast

Md Noh SM, Sheikh Abdul Kadir SH, Bannur ZM, Froemming GA, Abdul Hamid Hasani N, Mohd Nawawi H, et al.

Exp Eye Res, 2014 Oct;127:236-42.
PMID: 25139730 DOI: 10.1016/j.exer.2014.08.005

Anti-Vascular Endothelial Growth Factors (Anti-VEGF) agents have received recent interest as potential anti-fibrotic agents for their concurrent use with trabeculectomy. Preliminary cohort studies have revealed improved bleb morphology following trabeculectomy augmented with ranibizumab. The effects of this humanized monoclonal antibody on human Tenon's fibroblast (HTF), the key player of post trabeculectomy scar formation, are not fully understood. This study was conducted to understand the effects of ranibizumab on extracellular matrix production by HTF. The effect of ranibizumab on HTF proliferation and cell viability was determined using MTT assay (3-(4,5-dimethylthiazone-2-yl)-2,5-diphenyl tetrazolium). Ranibizumab at concentrations ranging from 0.01 to 0.5 mg/mL were administered for 24, 48 and 72 h in serum and serum free conditions. Supernatants and cell lysates from samples were assessed for collagen type 1 alpha 1 and fibronectin mRNA and protein level using quantitative real time polymerase chain reaction (qRT-PCR) and enzyme-linked immunosorbent assay (ELISA). After 48-h, ranibizumab at 0.5 mg/mL, significantly induced cell death under serum-free culture conditions (p cell viability in cultured HTF. From this study, we found that single application of ranibizumab is inadequate to induce the anti-fibrotic effects on HTF, suggesting the importance of adjunctive therapy. Further studies are underway to understand mechanism of actions of ranibizumab on HTF.

Matched MeSH terms: Cell Culture Techniques
Long-term evaluation of osteoarthritis sheep knee, treated with TGF-β3 and BMP-6 induced multipotent stem cells

Ude CC, Shamsul BS, Ng MH, Chen HC, Ohnmar H, Amaramalar SN, et al.

Exp Gerontol, 2018 04;104:43-51.
PMID: 29421350 DOI: 10.1016/j.exger.2018.01.020

BACKGROUND: Hyaline articular cartilage, which protects the bones of diarthrodial joints from forces associated with load bearing, frictions, and impacts has very limited capacities for self-repair. Over the years, the trend of treatments has shifted to regenerations and researchers have been on the quest for a lasting regeneration. We evaluated the treatment of osteoarthritis by chondrogenically induced ADSCs and BMSCs for a long time functional recovery.
METHODS: Osteoarthritis was induced at the right knee of sheep by complete resection of ACL and medial meniscus. Stem cells from sheep were induced to chondrogenic lineage. Test sheep received 5 mls single doses of 2 × 107 autologous PKH26-labelled ADSCs or BMSCs, while controls received basal medium. Functional recovery of the knees was evaluated via electromyography.
RESULTS: Induced ADSCs had 625, 255, 393, 908, 409, 157 and 1062 folds increases of collagen I, collagen II, aggrecan, SOX9, cartilage oligomeric protein, chondroadherin and fibromodullin compare to uninduced cells, while BMSCs had 702, 657, 321, 276, 337, 233 and 1163 respectively; p = .001. Immunocytochemistry was positive for these chondrogenic markers. 12 months post-treatment, controls scored 4 in most regions using ICRS, while the treated had 8; P = .001. Regenerated cartilages were positive to PKH26 and demonstrated the presence of condensing cartilages on haematoxylin and eosin; and Safranin O. OA degenerations caused significant amplitude shift from right to left hind limb. After treatments, controls persisted with significant decreases; while treated samples regained balance.
CONCLUSIONS: Both ADSCs and BMSCs had increased chondrogenic gene expressions using TGF-β3 and BMP-6. The treated knees had improved cartilage scores; PKH26 can provide elongated tracking, while EMG results revealed improved joint recoveries. These could be suitable therapies for osteoarthritis.

Matched MeSH terms: Cell Culture Techniques
Effects of IL-6, IL-10 and TGF-β on the expression of survivin and apoptosis in nasopharyngeal carcinoma TW01 cells

Poh YW, Gan SY, Tan EL

Exp Oncol, 2012 Jul;34(2):85-9.
PMID: 23013758

The aim of this study is to investigate whether IL-6, IL-10 and TGF-β are able to confer resistance to apoptosis in nasopharyngeal carcinoma cells by upregulating the expression of survivin.

Matched MeSH terms: Cell Culture Techniques
Transcriptional and physiological responses to inorganic nutrition in a tropical Pacific strain of Alexandrium minutum: Implications for nutrient uptakes and assimilation

Hii KS, Lim PT, Kon NF, Usup G, Gu H, Leaw CP

Gene, 2019 Aug 30;711:143950.
PMID: 31255736 DOI: 10.1016/j.gene.2019.143950

The marine dinoflagellate Alexandrium minutum is known to produce saxitoxins that cause paralytic shellfish poisoning in human worldwide through consumption of the contaminated shellfish mollusks. Despite numerous studies on the growth physiology and saxitoxin production of this species, the knowledge on the molecular basis of nutrient uptakes in relation to toxin production in this species is limited. In this study, relative expressions of the high-affinity transporter genes of nitrate, ammonium, and phosphate (AmNrt2, AmAmt1 and AmPiPT1) and the assimilation genes, nitrate reductase (AmNas), glutamine synthase (AmGSIII) and carbamoyl phosphate synthase (AmCPSII) from A. minutum were studied in batch clonal culture condition with two nitrogen sources (nitrate: NO3- or ammonium: NH4+) under different N:P ratios (high-P: N:P of 14 and 16, and low-P: N:P of 155). The expression of AmAmt1 was suppressed in excess NH4+-grown condition but was not observed in AmNrt2 and AmNas. Expressions of AmAmt1, AmNrt2, AmNas, AmGSIII, AmCPSII, and AmPiPT1 were high in P-deficient condition, showing that A. minutum is likely to take up nutrients for growth under P-stress condition. Conversely, relative expression of AmCPSII was incongruent with cell growth, but was well correlated with toxin quota, suggesting that the gene might involve in arginine metabolism and related toxin production pathway. The expression of AmGSIII is found coincided with higher toxin production and is believed to involve in mechanism to detoxify the cells from excess ammonium stress. The gene regulation observed in this study has provided better insights into the ecophysiology of A. minutum in relation to its adaptive strategies in unfavorable environments.

Matched MeSH terms: Batch Cell Culture Techniques/methods*
Valproic acid: growth inhibition of head and neck cancer by induction of terminal differentiation and senescence

Gan CP, Hamid S, Hor SY, Zain RB, Ismail SM, Wan Mustafa WM, et al.

Head Neck, 2012 Mar;34(3):344-53.
PMID: 21438066 DOI: 10.1002/hed.21734

There are limited studies on the effects of drugs that modulate epigenetic regulation for head and neck squamous cell carcinoma (HNSCC). This study determined the effect of valproic acid (VPA) on HNSCC.

Matched MeSH terms: Cell Culture Techniques
Cytotoxic and antioxidant effects of methoxylated stilbene analogues on HepG2 hepatoma and Chang liver cells: Implications for structure activity relationship

Hasiah AH, Ghazali AR, Weber JF, Velu S, Thomas NF, Inayat Hussain SH

Hum Exp Toxicol, 2011 Feb;30(2):138-44.
PMID: 20385705 DOI: 10.1177/0960327110368739

Stilbenes possess a variety of biological activities including chemopreventive activity. This study was conducted to evaluate the structural activity relationships of six methoxylated stilbene analogues with respect to their cytotoxic effects and antioxidant activities on HepG2 hepatoma and Chang liver cells. The cytotoxic and total antioxidant activities of six stilbene analogues were determined by MTT and Ferric Reducing Antioxidant Power (FRAP) assays, respectively. We found that the cis-methoxylated stilbene: (Z)-3,4,4'-trimethoxystilbene was the most potent and selective antiproliferative agent (IC₅₀ 89 µM) in HepG2 cells. For the total antioxidant activity, compounds possessing hydroxyl groups at the 4' position namely (E)-3-methoxy-4'-hydroxystilbene, (E)-3,5-dimethoxy-4'-hydroxystilbene (pterostilbene), (E)-4-methoxy-4'-hydroxystilbene showed the highest antioxidant activity. Structure activity relationship studies of these compounds demonstrated that the cytotoxic effect and antioxidant activities of the tested compounds in this study were structurally dependent.

Matched MeSH terms: Cell Culture Techniques
Identification of suitable culture condition for expansion and osteogenic differentiation of human bone marrow stem cells

Chowdhury SR, Ng MH, Hassan NS, Aminuddin BS, Ruszymah BH

Hum. Cell, 2012 Sep;25(3):69-77.
PMID: 22968953

This study was undertaken in order to identify the best culture strategy to expand and osteogenic differentiation of human bone marrow stem cells (hBMSCs) for subsequent bone tissue engineering. In this regard, the experiment was designed to evaluate whether it is feasible to bypass the expansion phase during hBMSCs differentiation towards osteogenic lineages by early induction, if not identification of suitable culture media for enhancement of hBMSCs expansion and osteogenic differentiation. It was found that introduction of osteogenic factors in alpha-minimum essential medium (αMEM) during expansion phase resulted in significant reduction of hBMSCs growth rate and osteogenic gene expressions. In an approach to identify suitable culture media, the growth and differentiation potential of hBMSCs were evaluated in αMEM, F12:DMEM (1:1; FD), and FD with growth factors. It was found that αMEM favors the expansion and osteogenic differentiation of hBMSCs compared to that in FD. However, supplementation of growth factors in FD, only during expansion phase, enhances the hBMSCs growth rate and significantly up-regulates the expression of CBFA-1 (the early markers of osteogenic differentiation) during expansion, and, other osteogenic genes at the end of induction compared to the cells in αMEM and FD. These results suggested that the expansion and differentiation phase of the hBMSCs should be separately and carefully timed. For bone tissue engineering, supplementation of growth factors in FD only during the expansion phase was sufficient to promote hBMSCs expansion and differentiation, and preferably the most efficient culture condition.

Matched MeSH terms: Cell Culture Techniques/methods*
Value of human amniotic epithelial cells in tissue engineering for cornea

Fatimah SS, Ng SL, Chua KH, Hayati AR, Tan AE, Tan GC

Hum. Cell, 2010 Nov;23(4):141-51.
PMID: 21166885 DOI: 10.1111/j.1749-0774.2010.00096.x

Human amniotic epithelial cells (hAECs) are potentially one of the key players in tissue engineering due to their easy availability. The aim of the present study was to develop an optimal isolation and transportation technique, as well as to determine the immunophenotype and epithelial gene expression of hAECs. Amnion was mechanically peeled off from the chorion and digested with trypsin-ethylenediaminetetraacetic acid. The isolated hAECs were cultured in medium containing 10 ng/mL epidermal growth factor until P4. The epithelial gene expression, cell surface antigen and protein expression of hAECs were analyzed by quantitative polymerase chain reaction, flow cytometry and immunocytochemistry. hAECs were also cultured in adipogenic, osteogenic and neurogenic induction media. The best cell yield of hAECs was seen in the digestion of 15 pieces of amnion (2 × 2 cm) and isolated 30 min after digestion with trypsin. F12:Dulbecco's modified eagle medium was the best medium for short term storage at 4 °C. hAECs expressed CD9, CD44, CD73 and CD90, and negligibly expressed CD31, CD34, CD45 and CD117. After serial passage, CK3, CK19 and involucrin gene expressions were upregulated, while p63, CK1 and CK14 gene expressions were downregulated. Sustained gene expressions of integrin β1 and CK18 were observed. At initial culture, these cells might have stem-like properties. However, they differentiated after serial passage. Nonetheless, hAECs have epithelial stem cell characteristics and have the potential to differentiate into corneal epithelial cells. Further investigations are still needed to elucidate the mechanism of differentiation involved and to optimize the culture condition for long term in vitro culture.

Matched MeSH terms: Cell Culture Techniques/methods*
Cell growth, cell-cycle progress, and antibody production in hybridoma cells cultivated under mild hypothermic conditions

Chong SL, Mou DG, Ali AM, Lim SH, Tey BT

Hybridoma (Larchmt), 2008 Apr;27(2):107-11.
PMID: 18642675

The effect of mild hypothermic (32 degrees C) conditions on cell growth, cell-cycle progress, and antibody production of hybridoma C2E7 cells was investigated in the present study. The growth of hybridoma cells was slower during the mild hypothermic condition compared to that at 37 degrees C; this led to about 10% decrease in maximum viable cell density and volumetric antibody productivity. However, under mild hypothermic growth conditions, the culture viability was substantially improved and the specific antibody productivity was enhanced compared to that at 37 degrees C. The average specific productivity for the entire batch culture at 32 degrees C is about 5% higher than that at 37 degrees C. Cell-cycle analysis data showed that there was no growth arrestment during the mild hypothermic growth of hybridoma cells. The G1-phase cells were increased, while the S-phase cells were decreased gradually as the culture time progressed. Further analysis showed that the specific antibody productivity of hybridoma cells was correlated to the fraction of S-phase cells.

Matched MeSH terms: Cell Culture Techniques
Comparison of different methods for the isolation of mesenchymal stem cells from human umbilical cord Wharton's jelly

Salehinejad P, Alitheen NB, Ali AM, Omar AR, Mohit M, Janzamin E, et al.

In Vitro Cell Dev Biol Anim, 2012 Feb;48(2):75-83.
PMID: 22274909 DOI: 10.1007/s11626-011-9480-x

Several techniques have been devised for the dissociation of tissues for primary culture. These techniques can affect the quantity and quality of the isolated cells. The aim of our study was to develop the most appropriate method for the isolation of human umbilical cord-derived mesenchymal (hUCM) cells. In the present study, we compared four methods for the isolation of hUCM cells: three enzymatic methods; collagenase/hyaluronidase/trypsin (CHT), collagenase/trypsin (CT) and trypsin (Trp), and an explant culture (Exp) method. The trypan blue dye exclusion test, the water-soluble tetrazolium salt-1 (WST-1) assay, flow cytometry, alkaline phosphatase activity and histochemical staining were used to evaluate the results of the different methods. The hUCM cells were successfully isolated by all methods but the isolation method used profoundly altered the cell number and proliferation capacity of the isolated cells. The cells were successfully differentiated into adipogenic and osteogenic lineages and alkaline phosphatase activity was detected in the hUCM cell colonies of all groups. Flow cytometry analysis revealed that CD44, CD73, CD90 and CD105 were expressed in all groups, while CD34 and CD45 were not expressed. The expression of C-kit in the enzymatic groups was higher than in the explant group, while the expression of Oct-4 was higher in the CT group compared to the other groups. We concluded that the collagenase/trypsin method of cell isolation yields a higher cell density than the others. These cells expressed a higher rate of pluripotent cell markers such as C-kit and Oct-4, while the explant method of cell isolation resulted in a higher cell proliferation rate and activity compared to the other methods.

Matched MeSH terms: Cell Culture Techniques
Development and partial characterization of new marine cell line from brain of Asian sea bass Lates calcarifer for virus isolation

Hasoon MF, Daud HM, Abdullah AA, Arshad SS, Bejo HM

In Vitro Cell Dev Biol Anim, 2011 Jan;47(1):16-25.
PMID: 21082288 DOI: 10.1007/s11626-010-9348-5

A new cell line, Asian sea bass brain (ASBB), was derived from the brain tissue of Asian sea bass Lates calcarifer. This cell line was maintained in Leibovitz L-15 media supplemented with 10% fetal bovine serum (FBS). The ASBB cell line was subcultured more than 60 times over a period of 15 mo. The ASBB cell line consists predominantly of fibroblastic-like cells and was able to grow at temperatures between 20°C and 30°C with an optimum temperature of 25°C. The growth rate of these cells increased as the proportion of FBS increased from 2% to 20% at 25°C with optimum growth at the concentrations of 10% or 15% FBS. Polymerase chain reaction products were obtained from ASBB cells and tissues of sea bass with primer sets of microsatellite markers of sea bass. An isolate of piscine nodavirus from juveniles of marine fish species tested positive by IQ2000 kit for viral nervous necrosis detection and was examined for its infectivity to a fish cell line of ASBB. A marine fish betanodavirus was tested to determine the susceptibility of this new cell line in comparison with commercial highly permissive SSN-1 cells. The ASBB cell line was found to be susceptible to nodavirus (RGNNV genotype), and the infection was confirmed by comparison cytopathic effect (CPE) with commercial SSN-1 and reverse transcriptase-polymerase chain reaction. A nodavirus was further elucidated by electron microscopy, and the virus tested was shown to induce CPE on ASBB cells with significant high titer. This suggests that the ASBB cell line has good potential for the isolation of fish viruses.

Matched MeSH terms: Cell Culture Techniques
Micromanipulation of culture niche permits long-term expansion of dental pulp stem cells--an economic and commercial angle

Govindasamy V, Ronald VS, Totey S, Din SB, Mustafa WM, Totey S, et al.

In Vitro Cell Dev Biol Anim, 2010 Oct;46(9):764-73.
PMID: 20725801 DOI: 10.1007/s11626-010-9332-0

Stem cells isolated from dental pulp possess the capacity for self-renewal and the potential for multi-lineage differentiation. However, dental pulp stem cells have different characteristics in terms of their culture conditions. The success of stem cells culture is governed by its micro-environmental niche. Therefore, we studied the effects of culture niche on long-term expansion of dental pulp stem cells in terms of cell morphology, growth kinetics, senescence pattern, cell surface marker expression differentiation capacity, and seeding plating density of dental pulp stem cells in four different, widely used media composition Among the various basal media tested, α-minimum essential media and knock out-minimum essential media supplemented with 10% fetal bovine serum were found to be the most optimal media composition in preserving the phenotypic characteristics and differentiation potential for prolonged periods as compared with DMEM-F12 and DMEM-LG. Plating density has been shown to affect overall yield. As a conclusion, the adoption of an appropriate culture system significantly improved cell yield, thus enabling the attainment of sufficient yields for therapeutic applications economizing in terms of cost of production and minimizing seeding cell density for maximum yield.

Matched MeSH terms: Cell Culture Techniques/methods*
Effect of supplementation of dermal fibroblasts conditioned medium on expansion of keratinocytes through enhancing attachment

Chowdhury SR, Aminuddin BS, Ruszymah BH

Indian J Exp Biol, 2012 May;50(5):332-9.
PMID: 22803323

In the present study in vitro expansion of human keratinocytes by supplementing dermal fibroblasts conditioned medium (DFCM) has been reported. Effect of two different DFCM acquired by culturing fibroblasts in keratinocyte-specific medium (defined keratinocytes serum free medium, DFCM-DKSFM) and fibroblast-specific serum free medium (F12: DMEM nutrient mix, DFCM-FD) have been compared. Growth kinetics of keratinocytes in terms of efficiency of cell attachment, expansion index, apparent specific growth rate and growth potential at the end of culture was evaluated in culture supplemented with DFCM-DKSFM and DFCM-FD in comparison with control i.e. DKSFM only. Results indicated that supplementation of DFCM caused significant increase in keratinocyte attachment. Efficiency of keratinocyte attachment in culture supplemented with bFCM-DKSFM was significantly higher compared to those cultured in DFCM-FD and DKSFM. In addition, the expansion index of keratinocytes in cultures supplemented with DFCM-DKSFM and DFCM-FD were 3.7 and 2.2 times higher than that of control condition even though the apparent growth rate and proliferative potential was found significantly lower. These results suggested that supplementation of DFCM enhanced expansion of keratinocyte by increasing efficiency of cell attachment, and DFCM-DKSFM provided suitable condition for in vitro expansion of keratinocytes compared to DFCM-FD and control condition.

Matched MeSH terms: Cell Culture Techniques
Fulltext Effect of Ganoderma lucidum polysaccharides on the growth of Bifidobacterium spp. as assessed using Real-time PCR

Yamin, S., Shuhaimi, M., Arbakariya, A., Khalilah, A. K., Anas, O., Yazid, A. M., et al.

International Food Research Journal, 2012;19(3):1199-1205.
MyJurnal

The use of component from Ganoderma lucidum as prebiotic source is interesting as the G. lucidum itself was known for more than a decade in the traditional Chinese medicine. In this work, Ganoderma lucidum crude polysaccharides (GLCP) and Polysaccharide-fraction number 2 (PF-2) were used as carbon sources in the fermentation with Bifidobacterium sp. The results showed the potential of prebiotic effect of the G. lucidum extract in batch-culture fermentation based on increment in the growth of bacteria used (0.4 – 1.5 log10 CFU/mL) after 18h fermentation. Fermentation was further done using faecal materials as bacterial inocula and bacterial growth changes were examined using real-time PCR. The results showed the ability of GLCP and PF-2 to support the growth of Bifidobacterium genus with 0.3 and 0.7 log10 cells/ml increased, respectively. Interestingly, Lactobacillus which is known as beneficial bacterial genus also showed growth increment with 0.7 and 1 log10 cells/ml increased. The competition for carbon sources thus inhibits the growth of potentially harmful genus, Salmonella (0.3 and 0.5 log10 cells/ml) in comparison to the control.

Matched MeSH terms: Batch Cell Culture Techniques

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