Journal: BMC Microbiology

Article Title: First human cell-based cultivation system for the syphilis spirochete Treponema pallidum

doi: 10.1186/s12866-026-04856-5

Figure Lengend Snippet: Human foreskin fibroblast cell lines support the growth of T. pallidum in vitro . A Human foreskin fibroblast cell lines (MoNa, HFF1, and HFFC) support the growth of T. pallidum in long-term, continuous, in vitro cultivation. While cultivation of strain SS14 was discontinued after 10 weeks, in vitro cultures of the DAL-1 strain have been ongoing for over one year. During this period, scheme of subcultures was optimized (up to 1:20). A booster passage (*), extending the standard 7-day subculture to a 14-day period with a cultivation medium exchange on day 7, was employed to enrich the treponemal culture during critical declines. In vitro cultivation was performed in triplicate (i.e., in three cultivation wells) and the mean values for each subculture are presented in graph. Source data from individual wells are shown in Supplementary Table . B Parallel cultivation for seven days ( n = 15) with defined DAL-1 inoculum (10 6 treponemes) validated the differences in growth support of individual foreskin cell lines. Treponemal growth on human foreskin fibroblast cells was slower compared to growth on rabbit epithelial Sf1Ep cells. Red bar, mean. C Human foreskin-based cultivation system supports the growth of various T. pallidum strains ( n = 8), from the Nichols-like as well as the SS14-like cluster. Note that human foreskin fibroblast cells were prepared as an equal mixture of three tested cell lines. Each T. pallidum strain was cultivated in a single in vitro well, representing a sole experimental replicate used for data acquisition

Article Snippet: Human foreskin fibroblasts HFF1 (SCRC-1041; ATCC) and HFFC (300715; Cytion) were purchased, while the third cell line (MoNa) was kindly provided by Dr. Vladimir Rotrekl (Masaryk University), and was originally obtained from the National Tissue Centre (Czech Republic).

Techniques: In Vitro

Journal: Molecular cancer research : MCR

Article Title: A Functional Precision Oncology Approach to Identify Treatment Strategies for Myxofibrosarcoma Patients

doi: 10.1158/1541-7786.MCR-21-0255

Figure Lengend Snippet: (A, B) Ex vivo drug testing of the novel patient derived cell model (WCM197) together with the OH931, NMFH-1 and HFF (benign human fibroblasts) using abemaciclib a second line CDK4/6-inhibitor and pralatrexate an antifolate inhibitor in a 6–8-point dilution dose response log scale to determine the IC50 values, highest dose was 10uM and lowest dose was 4.7 pM with an assay time of 96 hours. Greater cytostatic and cytotoxic effect was seen in the cells with the CDKN2A/B and MTAP null status (WCM197 and OH931) compared to CDKN2A/B and MTAP wild type status (NMFH-1 and HFF). (C) The combination of pralatrexate (IC30) and abemaciclib killed all our patient cells within 48 hours compared to the vehicle and abemaciclib and pralatrexate mono-therapy. (D) WCM197 sarco-sphere formation is completely inhibited with the combination of abemaciclib and pralatrexate. (E) Drug combination of abemaciclib and pralatrexate causes massive cell swellings and cytoplasmatic vacuole formation. (F) Cleaved caspase 3 and cleaved PARP at 24, 48 and 72 hours were detected with the combination of abemaciclib and pralatrexate indicating the induction of cell death in the tumor cells after treatment.

Article Snippet: The Human Foreskin Fibroblast (HFF) cells were provided by Dr. Carla Grandori, SEngine Precision Medicine, Seattle, WA, USA as a control specimen for high throughput drug screening ( 22 ).

Techniques: Ex Vivo, Derivative Assay

Journal: Molecular cancer research : MCR

Article Title: A Functional Precision Oncology Approach to Identify Treatment Strategies for Myxofibrosarcoma Patients

doi: 10.1158/1541-7786.MCR-21-0255

Figure Lengend Snippet: (A) High throughput drug screening of all 4 cell strains (HFF, OH931, WCM197, NMFH-1) shown as a heat map (blue is sensitive to red resistant). RB1 wt and CDKN2A/B/MTAP null cells (WCM197 and OH931) show higher sensitivity to CDK4/6 inhibititors such as abemaciclib (indicated by the yellow arrows) and the antifolate pralatrexate and methotrexate (indicated by the green arrows) compared to the CDKN2A/B/MTAP wild type lines (NMFH-1 and HFF). Selective sensitivity to PLK-1 inhibitors was detected in WCM197 and OH931 cells compared to the NMFH-1 and the control line HFF, shown in orange arrows. (B) Graphs show the response of the sarcoma cells to each compound in the library as area under the curve (AUC) compared to the benign human foreskin fibroblast line (HFF) as a normal control. (C-D) Ex vivo drug validation of the two PLK-1 inhibitors (volasertib and rigosertib) confirms high sensitivity for WCM197 and OH931 cells. (E, F) WCM197 3D sarco-spheres are completely inhibited in growth under volasertib treatment and show cell death indicated by cleaved caspase 3, PARP and cleaved PARP. Cell death was monitored with cleaved caspase 3 and cleaved PARP over 96 hours with a peak at 24–72 hours.

Article Snippet: The Human Foreskin Fibroblast (HFF) cells were provided by Dr. Carla Grandori, SEngine Precision Medicine, Seattle, WA, USA as a control specimen for high throughput drug screening ( 22 ).

Techniques: High Throughput Screening Assay, Drug discovery, Control, Ex Vivo, Biomarker Discovery

Journal: mBio

Article Title: Accessory Gene Regulator-1 Locus Is Essential for Virulence and Pathogenesis of Clostridium difficile

doi: 10.1128/mBio.01237-16

Figure Lengend Snippet: Culture supernatant fluids collected from the agrB1D1 mutants are not cytotoxic to human foreskin fibroblast cells. Strain 630 and R20291 agrB1D1 and agrB2D2 mutants were cultured for 48 h in BHI medium anaerobically at 37°C. The culture supernatant fluids were collected, filter sterilized with a 0.45-µm filter, and examined for cytotoxicity with the Bartels Clostridium difficile cytotoxicity assay kit (Trinity Biotech, Jamestown, NY). The culture fluids were incubated with the fibroblast cells for 24 h and observed under a microscope for cytotoxic effects. Images were taken with an EVOS XL microscope (Advanced Microscopy Group) at ×20 magnification. Panels: A, a representative image of fibroblast cells cultured in growth medium only; B, wild-type 630; C, 630 agrB1D1 mutant; D, 630 complemented agrB1D1 mutant; E, wild-type R20291; F, R20291 agrB1D1 mutant; G, complemented R20291 agrB1D1 mutant; H, R20291 agrB2D2 mutant.

Article Snippet: Ready-made human foreskin fibroblast cells (Trinity Biotech, Jamestown, NY) in 150 μl of medium were treated with 50 μl of culture supernatant fluids from the wild-type and agr mutant strains and incubated for 48 h at 37°C.

Techniques: Cell Culture, Cytotoxicity Assay, Incubation, Microscopy, Mutagenesis

Journal: mBio

Article Title: Accessory Gene Regulator-1 Locus Is Essential for Virulence and Pathogenesis of Clostridium difficile

doi: 10.1128/mBio.01237-16

Figure Lengend Snippet: Culture supernatants from C. difficile agrB1D1 mutants have no effect on cell viability. Strain 630 (A) and R20291 (B) agrB1D1 and agrB2D2 mutants were cultured for 48 h in BHI medium anaerobically at 37°C. The culture supernatant fluids were collected, filter sterilized with a 0.45-µm filter, and tested for their effect on cell viability. Briefly, ready-made fibroblast cells (Trinity Biotech, Jamestown, NY) were treated with the culture supernatant fluids and incubated for 48 h at 37°C aerobically. Following the incubation period, cell viability was examined with the CellTiter 96 nonradioactive cell proliferation assay (Promega, Madison, WI) in accordance with the instructions provided by the manufacturer. agrB1D1 Mut, agrB1D1 deletion mutant; agrB1D1 Mut+Comp, agrB1D1 mutant complemented with a plasmid bearing the wild-type agrB1D1 locus; agrB2D2 Mut, R20291 agrB2D2 deletion mutant. The differences between the amounts of formazan produced by fibroblast cells treated with supernatants from the agrB1D1 mutants and wild-type strains, the complemented agrB1D1 mutants, and the R20291 agrB2D2 mutant were significant ( P = 0.0012). Error bars represent the standard deviations of three independent experiments.

Article Snippet: Ready-made human foreskin fibroblast cells (Trinity Biotech, Jamestown, NY) in 150 μl of medium were treated with 50 μl of culture supernatant fluids from the wild-type and agr mutant strains and incubated for 48 h at 37°C.

Techniques: Cell Culture, Incubation, Proliferation Assay, Mutagenesis, Plasmid Preparation, Produced

Journal: International Journal for Parasitology: Drugs and Drug Resistance

Article Title: In vitro activity, safety and in vivo efficacy of the novel bumped kinase inhibitor BKI-1748 in non-pregnant and pregnant mice experimentally infected with Neospora caninum tachyzoites and Toxoplasma gondii oocysts

doi: 10.1016/j.ijpddr.2021.05.001

Figure Lengend Snippet: Structure and in vitro activities of BKI-1748 against N. caninum (Nc-β-gal) and T. gondii (Tg-β-gal). (A) molecular structure of BKI-1748, MW = molecular weight; EC 50 = concentration of half maximal proliferation inhibition; *toxicity levels against human foreskin fibroblasts (HFF) and zebrafish embryos were determined previously [22]. (B,C) Dose-response curves for Neospora and Toxoplasma tachyzoites grown in HFF.

Article Snippet: Human foreskin fibroblasts (HFF; PCS-201-010TM) and BALB/c dermal fibroblasts (CELLNTEC AG, Bern, Switzerland) were maintained as described ( ).

Techniques: In Vitro, Molecular Weight, Concentration Assay, Inhibition

Journal: International Journal for Parasitology: Drugs and Drug Resistance

Article Title: In vitro activity, safety and in vivo efficacy of the novel bumped kinase inhibitor BKI-1748 in non-pregnant and pregnant mice experimentally infected with Neospora caninum tachyzoites and Toxoplasma gondii oocysts

doi: 10.1016/j.ijpddr.2021.05.001

Figure Lengend Snippet: Litter size, parasite burden, neonatal and postnatal mortality rates of N. caninum infected mice treated with BKI-1748.

Article Snippet: Human foreskin fibroblasts (HFF; PCS-201-010TM) and BALB/c dermal fibroblasts (CELLNTEC AG, Bern, Switzerland) were maintained as described ( ).

Techniques: Infection