ABSTRACT
This study aimed to investigate the effect of in vivo pH-responsive doxorubicin (DOX) release and the targetability of pilot molecules in folic acid (FA)-modified micelles using a pharmacokinetic-pharmacodynamic (PK-PD) model. The time profiles of intratumoral DOX concentrations in Walker256 tumor-bearing rats were monitored using a microdialysis probe, followed by compartmental analysis, to evaluate intratumoral tissue pharmacokinetics. Maximal DOX was released from micelles 350 min after the administration of pH-responsive DOX-releasing micelles. However, FA modification of the micelles shortened the time to peak drug concentration to 150 min. Additionally, FA modification resulted in a 27-fold increase in the tumor inflow rate constant. Walker256 tumor-bearing rats were subsequently treated with DOX, pH-responsive DOX-releasing micelles, and pH-responsive DOX-releasing FA-modified micelles to monitor the tumor growth-time profiles. An intratumoral threshold concentration of DOX (55-64 ng/g tumor) was introduced into the drug efficacy compartment to construct a PD model, followed by PK-PD analysis of the tumor growth-time profiles. Similar results of threshold concentration and drug potency of DOX were obtained across all three formulations. Cell proliferation was delayed as the drug delivery ability of DOX was improved. The PK model, which was developed using the microdialysis method, revealed the intratumoral pH-responsive DOX distribution profiles. This facilitated the estimation of intratumoral PK parameters. The PD model with threshold concentrations contributed to the estimation of PD parameters in the three formulations, with consistent mechanisms observed. We believe that our PK-PD model can objectively assess the contributions of pH-responsive release ability and pilot molecule targetability to pharmacological effects.
Subject(s)
Doxorubicin , Folic Acid , Micelles , Doxorubicin/pharmacokinetics , Doxorubicin/pharmacology , Doxorubicin/administration & dosage , Doxorubicin/chemistry , Animals , Rats , Hydrogen-Ion Concentration , Folic Acid/chemistry , Folic Acid/pharmacokinetics , Drug Liberation , Cell Proliferation/drug effects , Drug Delivery Systems/methods , Cell Line, Tumor , Drug Carriers/chemistry , Female , Rats, Wistar , Humans , Antibiotics, Antineoplastic/pharmacokinetics , Antibiotics, Antineoplastic/administration & dosage , Antibiotics, Antineoplastic/pharmacologyABSTRACT
Translin, a ubiquitous RNA/DNA-binding protein that forms a hetero-octamer together with Translin-associated factor X (TRAX), possesses endoribonuclease activity and plays a physiological role in restricting the size and differentiation of mesenchymal precursor cells. However, the precise role of Translin in epithelial cells remains unclear. Here, we show evidence that Translin restricts the growth of pubertal mammary epithelial cells. The mammary epithelia of Translin-null females exhibited retarded growth before puberty, but highly enhanced growth and DNA synthesis with increased ramification after the onset of puberty. Primary cultures of Translin-null mammary epithelial cells showed augmented DNA synthesis in a ligand-independent and ligand-enhanced manner. Translin-null ovariectomized mice implanted with slow-release estrogen pellets showed enhanced length and ramification of the mammary glands. Mammary epithelial growth was also observed in ovariectomized Translin-null mice implanted with placebo pellets. Luciferase reporter assays using embryonic fibroblasts from Translin-null mice showed unaltered estrogen receptor α function. These results indicate that Translin plays a physiological role in restricting intrinsic growth, beyond mesenchymal cells, of pubertal mammary epithelial cells.
Subject(s)
DNA-Binding Proteins/metabolism , Epithelial Cells/cytology , Mammary Glands, Animal/growth & development , RNA-Binding Proteins/metabolism , Animals , Cell Proliferation , Cells, Cultured , DNA Replication , DNA-Binding Proteins/genetics , Epithelial Cells/metabolism , Female , Gene Deletion , Mammary Glands, Animal/cytology , Mammary Glands, Animal/metabolism , Mice , Mice, Inbred C57BL , Mice, Knockout , RNA-Binding Proteins/genetics , Sexual MaturationABSTRACT
Translin, a highly conserved DNA/RNA binding protein that forms a hetero-octamer together with Translin-associated factor X (TRAX), possesses a broad variety of functions, including RNA processing and DNA repair. Recent studies have reported that Translin is involved in mesenchymal cell physiology. Thus, here we analyzed the intrinsic role of Translin in mesenchymal cell proliferation and differentiation. Translin-deficient E11.5 mouse embryonic fibroblasts showed enhanced growth. Translin-deficient bone marrow-derived mesenchymal stem cells showed substantial expansion in vivo and enhanced proliferation in vitro. These cells also showed enhanced osteogenic and adipocytic differentiation. Histological analyses showed adipocytic hypertrophy in various adipose tissues. Translin knockout did not affect the growth of subcutaneous white adipose tissue-derived stem cells, but enhanced adipocytic differentiation was observed in vitro. Contrary to previous reports, in vitro-fertilized Translin-null mice were not runted and exhibited normal metabolic homeostasis, indicating the fragility of these mice to environmental conditions. Together, these data suggest that Translin plays an intrinsic role in restricting mesenchymal cell proliferation and differentiation.
Subject(s)
Bone Marrow Cells/cytology , DNA-Binding Proteins/metabolism , Mesenchymal Stem Cells/cytology , RNA-Binding Proteins/metabolism , Adipose Tissue/cytology , Animals , Cell Differentiation , Cell Proliferation , Female , Fibroblasts/cytology , Fibroblasts/metabolism , Male , Mice , Mice, Inbred C57BL , Mice, Knockout , OsteogenesisABSTRACT
The expression of extracellular matrix protein periostin (POSTN) was attenuated in Med1(-/-) mouse embryonic fibroblasts (MEFs), which exhibited a decreased capability to support hematopoietic progenitor cells (HPCs) in vitro. When bone marrow (BM) cells were cocultured with mitomycin C-treated Med1(+/+) MEFs, or OP-9 or MS-5 BM stromal cells, in the presence of anti-POSTN antibody, the growth of BM cells and number of long-term culture-initiating cells (LTC-ICs) were attenuated. When BM cells were cocultured with Med1(-/-) MEFs in the presence of recombinant POSTN, the growth of BM cells and the number of LTC-ICs were restored. Moreover, antibody-mediated blockage of stromal cells-derived POSTN markedly reduced the growth and cobblestone formation, a leukemic stem cell feature, of stromal cell-dependent MB-1 myeloblastoma cells. POSTN was expressed both in BM cells and variably in different BM stromal cells. Expression in the latter cells was increased by physical interaction with hematopoietic cells. The receptor for POSTN, integrin αvß3, was expressed abundantly in BM stromal cells. The addition of recombinant POSTN to BM stromal cells induced intracellular signaling downstream of integrin αvß3. These results suggest that stromal cell POSTN supports both normal HPCs and leukemia-initiating cells in vitro, at least in part, indirectly by acting on stromal cells in an autocrine or paracrine manner.
