Journal: PLoS ONE

Article Title: Exclusion of Exon 2 Is a Common mRNA Splice Variant of Primate Telomerase Reverse Transcriptases

doi: 10.1371/journal.pone.0048016

Figure Lengend Snippet: A. A phylogenetic tree of TERT mRNA coding regions was created with the Jalview algorithm using percent identity neighbor joining. A genome sequence is currently unavailable for Bonobo, but is in the same genus ( Pan ) as chimpanzee. B. RT-PCR of TERT exons 1–3 of Monkey (Cos-7 cells), Mouse (NIH3T3), Rat (NRK), Dog (D17), Orangutan ( Pongo pygmaeus ) and Bonobo ( Pan paniscus ). An arrow indicates wild type bands and an asterisk indicates DEL[e2]. PCR reactions shown are representative of at least 2 trials.

Article Snippet: A431 (ATCC CRL-1555; epidermoid carcinoma), HeLa (ATCC CCL-2; cervical epithelial adenocarcinoma), HEK293 (ATCC CRL-1573; adenovirus 5 transformed embryonic kidney cells), NRK (ATCC CRL-6509; normal rat kidney epithelial cells), D17 (ATCC CCL-183; dog osteosarcoma lung metastasis) and COS-7 (ATCC CRL-1651; SV40 transformed African green monkey kidney fibroblast cells) cells were cultured in DMEM (Gibco) with 10% FBS and 1% antibiotics.

Techniques: Sequencing, Reverse Transcription Polymerase Chain Reaction

Journal: BMC Veterinary Research

Article Title: The effects of taurolidine alone and in combination with doxorubicin or carboplatin in canine osteosarcoma in vitro

doi: 10.1186/1746-6148-9-15

Figure Lengend Snippet: Effects of doxorubicin or carboplatin ± taurolidine on proliferation of OS cell lines. Three OS cell lines were incubated for 72 hrs in increasing concentrations of doxorubicin A ) or carboplatin B ) in the presence or absence of taurolidine and cytotoxic effects were evaluated using an MTS assay. T0 indicates no taurolidine while T20 and T100 indicate 20 μM and 100 μM taurolidine respectively. Error bars indicate SD of 4 replicate wells. In some cases error bars are too small to be visible. * indicates a synergistic interaction (see methods). Data are presented as percent difference from time 0 on the y-axis (see methods and results). Photomicrographs of D17 cells after 6 hr incubation with 100 nM doxorubicin C ) or with 100 nM doxorubicin plus 20 μM taurolidine. D ) Cytoplasmic contraction is readily appreciated in the cells exposed to the drug combination and represents an early apoptotic effect.

Article Snippet: They included canine D17 (ATCC CCL183), Clone 4, developed in our laboratory [ ], COS [ ], and HMPOS [ ], and the human cell line SAOS-2 (ATCC HTB-85) which was included to establish consistency with reports using human cell lines.

Techniques: Incubation, MTS Assay

Journal: BMC Veterinary Research

Article Title: The effects of taurolidine alone and in combination with doxorubicin or carboplatin in canine osteosarcoma in vitro

doi: 10.1186/1746-6148-9-15

Figure Lengend Snippet: Mechanisms of t aurolidine-induced cytotoxicity are cell line and concentration dependent. Binding of annexin-V-biotin-FITC and propidium iodide (PI) to taurolidine-treated D17 OS cells was determined by 2-color flow cytometry. A ) Data are displayed as dot plots and data points in the lower right quadrant (i.e., annexin-V positive) represent early apoptotic cells while data points in the upper left quadrant represent necrotic cells and points in the upper right quadrant (dual annexin-V/PI positive) represent cells in late apoptosis and necrosis. B ) The release of nucleosomes as an indicator of apoptosis following taurolidine treatment of D17 OS cells was determined at two time points. Data are expressed as percent of apoptosis values measured in untreated cells at the same time points. The higher taurolidine concentration (250 μM) was less effective at inducing apoptotic changes in this assay indicating cell death occurred by other mechanisms. T125, T250 = 125 μM and 250 μM taurolidine respectively. * indicates different from 125 μM at same time point (p < .01). Error bars indicate SD. C ) Western blots show degradation of PARP as an indirect indicator of taurolidine-activated caspase activity in COS and HMPOS cells. Cells incubated 12 hrs in taurolidine as described in methods. No loading control was available for the D17 blot because it was an immunoprecipitation. D ) These proliferation assays show the caspase inhibitor (CI) Z-VAD-FMK protects HMPOS but not D17 OSA cells from caspase-dependent cell death at low concentrations of taurolidine, consistent with PARP cleavage seen in 3C. * indicates intercept is different between treated and untreated conditions (P < .05).

Article Snippet: They included canine D17 (ATCC CCL183), Clone 4, developed in our laboratory [ ], COS [ ], and HMPOS [ ], and the human cell line SAOS-2 (ATCC HTB-85) which was included to establish consistency with reports using human cell lines.

Techniques: Concentration Assay, Binding Assay, Flow Cytometry, Western Blot, Activity Assay, Incubation, Immunoprecipitation

Journal: BMC Veterinary Research

Article Title: The effects of taurolidine alone and in combination with doxorubicin or carboplatin in canine osteosarcoma in vitro

doi: 10.1186/1746-6148-9-15

Figure Lengend Snippet: IC50s for human and canine osteosarcoma cell lines treated with taurolidine or doxorubicin (inhibition curves shown in Figure 1 )

Article Snippet: They included canine D17 (ATCC CCL183), Clone 4, developed in our laboratory [ ], COS [ ], and HMPOS [ ], and the human cell line SAOS-2 (ATCC HTB-85) which was included to establish consistency with reports using human cell lines.

Techniques: Inhibition

Journal: BMC Veterinary Research

Article Title: The effects of taurolidine alone and in combination with doxorubicin or carboplatin in canine osteosarcoma in vitro

doi: 10.1186/1746-6148-9-15

Figure Lengend Snippet: IC50s for human and canine osteosarcoma cell lines treated with doxorubicin or carboplatin ± taurolidine (inhibition curves shown in Figure 2 A and 2 B)

Article Snippet: They included canine D17 (ATCC CCL183), Clone 4, developed in our laboratory [ ], COS [ ], and HMPOS [ ], and the human cell line SAOS-2 (ATCC HTB-85) which was included to establish consistency with reports using human cell lines.

Techniques: Inhibition

Journal: BMC Veterinary Research

Article Title: The effects of taurolidine alone and in combination with doxorubicin or carboplatin in canine osteosarcoma in vitro

doi: 10.1186/1746-6148-9-15

Figure Lengend Snippet: p53 protein accumulation in untreated osteosarcoma cells. Representative photomicrographs of cells probed with an antibody against p53 protein show strong accumulation in HMPOS cells, moderate staining in Clone-4 and none in the D17 and COS cell lines. The signal that can be observed in D17 and COS cells is the counterstain hematoxylin (see methods).

Article Snippet: They included canine D17 (ATCC CCL183), Clone 4, developed in our laboratory [ ], COS [ ], and HMPOS [ ], and the human cell line SAOS-2 (ATCC HTB-85) which was included to establish consistency with reports using human cell lines.

Techniques: Staining

Journal: BMC Veterinary Research

Article Title: The effects of taurolidine alone and in combination with doxorubicin or carboplatin in canine osteosarcoma in vitro

doi: 10.1186/1746-6148-9-15

Figure Lengend Snippet: Immunocytochemistry scores for p53 protein accumulation. 0 indicates no detection and +++ indicates strong staining in all cells

Article Snippet: They included canine D17 (ATCC CCL183), Clone 4, developed in our laboratory [ ], COS [ ], and HMPOS [ ], and the human cell line SAOS-2 (ATCC HTB-85) which was included to establish consistency with reports using human cell lines.

Techniques: Immunocytochemistry, Staining

Journal: BMC Veterinary Research

Article Title: The effects of taurolidine alone and in combination with doxorubicin or carboplatin in canine osteosarcoma in vitro

doi: 10.1186/1746-6148-9-15

Figure Lengend Snippet: Effects of taurolidine on p53 protein expression. Western blots showing the effects of taurolidine exposure on this cell cycle regulatory protein is concentration and cell line dependent. The taurolidine-induced appearance of p53 protein in D17 cells suggests these cells proceed through a p53-induced apoptotic pathway when exposed to 125 uM taurolidine. In HMPOS cells, with apparent mutant p53, p53 was down-regulated suggesting cell death may occur through different pathways in this cell line. Clone 4 and COS cells did not show any p53 expression at any concentration of taurolidine tested (data not shown).

Article Snippet: They included canine D17 (ATCC CCL183), Clone 4, developed in our laboratory [ ], COS [ ], and HMPOS [ ], and the human cell line SAOS-2 (ATCC HTB-85) which was included to establish consistency with reports using human cell lines.

Techniques: Expressing, Western Blot, Concentration Assay, Mutagenesis