Background Standard chemotherapy with taxanes, such as paclitaxel (PTX), remains the mainstay of systemic treatment of triple-negative breast cancer. Nanotechnology-based formulations have gradually replaced PTX injection and are widely used in China. However, no studies have compared the colloidal stability, antitumor efficacy, and safety of commercial PTX nanoformulations. Additionally, the desire to evaluate preclinical antitumor efficacy in human-derived tumor cells led to the widespread application of immunodeficient mouse models that likely contributed to the neglect of nanomedicines-immune system interactions. The present study investigated the colloidal stability, antitumor efficacy and safety, and nanomedicines-host immune system interactions of PTX nanoformulations. A further comparative analysis was performed to evaluate the clinical potential. Results Compared with liposome, PTX emulsion and PTX nanoparticle exhibited favorable colloidal stability. PTX emulsion was superior in inducing apoptosis and had a more pronounced inhibitory effect on 4T1-tumor spheroids compared with PTX liposome and PTX nanoparticle. Although PTX emulsion exhibited superior in vitro antitumor effect, no significant differences in the in vivo antitumor efficacy were found among the three types of PTX nanoformulations in an immunocompetent orthotopic 4T1 murine triple-negative breast cancer model. All PTX nanoformulations at maximum tolerated dose (MTD) induced lymphopenia and immunosuppression, as evidenced by the reduction of T cell subpopulations and inhibition of the dendritic cells maturation. Conclusions The MTD PTX nanomedicines-induced lymphopenia and immunosuppression may weaken the lymphocyte-mediated antitumor cellular immune response and partly account for the lack of differences in the in vivo antitumor outcomes of PTX nanoformulations. Understanding of what impacts PTX nanomedicines has on the immune system may be critical to improve the design and conduct of translational research of PTX nanomedicines in monotherapy or combination therapy with immunotherapy. Graphic abstract
This study developed somatic embryogenesis protocols for Picea pungens (Engelm), an important ornamental species, including initiation, proliferation, maturation, germination, and acclimation. Somatic embryogenic tissues were induced from mature zygotic embryos of five families, with a frequency of $$\ge $$ ≥ 22% for each. Embryogenic tissues (ET) from 13 clones of three families were proliferated for one week, achieving an average rate of 179.1%. The ET of 38 clones of three families were cultured in maturation medium for six weeks; 188 mature embryos on average were counted per gram ET cultured, of which $$\ge $$ ≥ 81.1% appeared normal, and each clone developed at least 28 normally matured embryos. A total of 69.9% or more of cotyledonary somatic embryos germinated normally and developed into normal emblings. The experiment of transplanting the emblings into a greenhouse had an average survival rate of 68.5%. Considerable variation among and within families during initiation and proliferation was observed, but this variation decreased in the maturation and germination. Changing the concentration of plant growth regulator of the initiation medium did not significantly change the initiation frequency. We recommend incorporating these protocols into the current Picea pungens practical programs, although further research is essential to increase efficiencies and reduce cost.
Picea pungens (Engelm.), known for its blue-green needles, has become a likable ornamental species in northeast China since 2000. Nonetheless, a lack of propagation methods that can maintain genetic fidelity and develop seedlings at a large scale prevents the further expansion of the species. Somatic embryogenesis (SE), paired with cryopreservation technologies, may provide a valid alternative. Picea pungens SE is not new, but its practical application has been limited due to low efficiencies in SE initiation and maturation as well as a lack of effective cryopreservation technology. In this study, experiments were carried out to overcome the limitations by modifying culture media. For initiation, the efficiency was enhanced by adjusting concentrations of 2.4-dichlorophenoxy acetic acid (2,4-D), 6-benzyl amino–purine (6-BA) or sucrose supplemented to the induction medium. The concentrations of 4.0 mg/L 2,4-D, 2 mg/L 6-BA, and 5 to 10 g/L sucrose were found optimal in maximizing initiation efficiency. For maturation, the efficiency, expressed as the number of mature somatic embryos per gram of fresh mass cultured (E/gFM), varied greatly with the choices of the basal medium and concentration of abscisic acid (ABA) of the maturation medium. Based on our results, the judicial choices were using the DCR medium as the basal medium and 10 mg/L ABA. The maturation efficiency could also be improved by adjusting the maturation medium’s osmotic pressure by manipulating the concentrations of carbohydrate and Gelrite and culture density. While the maturation medium, using sucrose as carbohydrate source or supplemented with a low (<8 g/L) Gelrite concentration, facilitated maturation, optimal selections were truly genotype-dependent. Our results also suggest that, while the optimal culture density varied with genotype, in general it is needless to culture more than 100 mg embryogenesis tissues per dish (size: 10 × 1.5 cm). Based on this study, the optimum pretreatment for embryogenesis tissue cryopreservation was culturing the tissues on the proliferation medium with 0.4 mol/L sorbitol for 24 h, followed by treatment with 5% Dimethyl sulfoxide. This study significantly improved the initiation (achieved a frequency of 0.56) and embryo maturation efficiencies (achieved 1030 E/gFM) and established an effective preculturing protocol for cryopreservation (recovered 1354 E/gFM) for the species. The protocols developed here, paired with the available ones for other SE steps in the literature, form a well-refined SE technology intended for commercial application to Picea pungens.
Embryogenic tissue (ET) is important for genetic modification and plant re-generation. The proliferation ability and vigor of ET are crucial for plant propagation via somatic embryogenesis. In this study, ET was induced from mature zygotic embryos in blue spruce (Picea pungens Engelm.). There were significant differences in ET induction between two provenances, i.e. 78.8 ± 12.5% and 62.50 ± 12.8% respectively. Effects of 2,4-Dichlorophenoxy acetic acid (2,4-D), 6-Benzyl amino-purine (6-BA) and/or sucrose on ET proliferation and somatic embryo (SE) maturation were further investigated with four cell lines. The highest ET proliferation rate reached 1473.7 ± 556.0% biweekly. Concentrations of 2,4-D or 6-BA applied at tissue proliferation stage impacted SE maturation among the cell lines, whereas sucrose showed less effects. The highest rate, 408 ± 230 mature SEs/g FW, was achieved in SE maturation cultures. This research demonstrated that the culture conditions, i.e. the specific concentrations of 2,4-D and BA, at ET proliferation stage affected not only ET growth, but also the quality of ET for SE maturation. This study revealed the necessity and benefit in developing both the general and the genotype-specific protocols for efficient production of mature SEs, or somatic plants in blue spruce.
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