ABSTRACT
BACKGROUND/AIMS: Acute kidney injury (AKI) is a common clinical condition that can lead to chronic kidney failure. Although mesenchymal stem cell-derived extracellular vesicles (MSC EVs) are regarded as a potent AKI treatment, the mechanisms underlying their beneficial effects remain unclear. Oct-4 may play an important role in tissue injury repair. We thus hypothesized that oct-4 overexpression might enhance the therapeutic effects of MSC EVs in AKI treatment. METHODS: Renal tubular epithelial cells were cultured in a low oxygen environment, then cocultured with MSC EVs or control medium for 48 h. BrdU and transferase-mediated dUTP nick-end labeling (TUNEL) staining were used to assess cell proliferation and apoptosis. Mice subjected to ischemia reperfusion were randomly divided into 4 groups, then injected with either phosphate-buffered saline (vehicle), EVs, EVs overexpressing oct-4 (EVs+Oct-4), and EVs not expressing Oct-4 (EVs-Oct-4). Blood creatinine (CREA) and urine nitrone levels were assessed 48 h and 2 weeks after injection. After ischemia reperfusion, renal tissues from each group were stained with TUNEL and proliferating cell nuclear antigen (PCNA) to determine the degree of apoptosis and proliferation. Masson trichrome staining was used to evaluate renal fibrosis progression. Snail gene expression was assessed using polymerase chain reaction (PCR). RESULTS: At 48 h after hypoxic treatment, TUNEL and BrdU staining indicated that the EVs+Oct-4 group had the least apoptosis and the most proliferation, respectively. Treatment with EVs overexpressing Oct-4 significantly decreased serum Crea and blood urea nitrogen levels and rescued kidney fibrosis, as indicated by the low proportion of Masson staining, high number of PCNA-positive cells, and low number of TUNEL-positive cells. PCR analysis indicated that Snail was most upregulated in the vehicle group and least upregulated in the EVs+Oct-4 group. CONCLUSIONS: MSC EVs had a pronounced therapeutic effect on ischemic reperfusion injury-related AKI, and Oct-4 overexpression enhanced these therapeutic effects. Our results may inspire a new direction for AKI treatment with MSC EVs.
Subject(s)
Acute Kidney Injury/therapy , Mesenchymal Stem Cell Transplantation/methods , Mesenchymal Stem Cells/metabolism , Octamer Transcription Factor-3/genetics , Acute Kidney Injury/pathology , Animals , Disease Models, Animal , Extracellular Vesicles/metabolism , Humans , Immunohistochemistry , Male , Mesenchymal Stem Cells/cytology , Mice , Mice, Inbred C57BL , Octamer Transcription Factor-3/biosynthesis , Random AllocationABSTRACT
The reprogramming of fibroblasts to induced pluripotent stem cells raises the possibility that a somatic cell can be reprogrammed to an alternative, differentiated fate without first becoming a stem/progenitor cell. Recent work has shown that fibroblasts can be reprogrammed to other, terminally differentiated cells with a combination of several transcription factors. Here, we report that a combination of four developmental transcription factors; GATA4, SF-1, NGFI-B, and COUP TF2; efficiently reprogrammed human foreskin fibroblasts into functional induced Leydig-like cells (iLCs). The iLCs expressed Leydig-specific markers and secreted testosterone in vitro. We found that GATA4 and SF-1 were particularly critical for Leydig-specific markers expression and that GATA4, SF-1, and NGFI-B were necessary to generate functional iLCs that secreted testosterone. These findings demonstrate that fibroblasts can be directly converted into iLCs with a few, defined factors and may provide insight into potential therapies to treat testosterone deficiency.
ABSTRACT
A novel gene (aga4436), encoding a potential agarase of 456 amino acids, was identified in the genome of deep-sea bacterium Flammeovirga sp. OC4. Aga4436 belongs to the glycoside hydrolase 16 ß-agarase family. Aga4436 was expressed in Escherichia coli as a fusion protein and purified. Recombinant Aga4436 showed an optimum agarase activity at 50-55 °C and pH 6.5, with a wide active range of temperatures (30-80 °C) and pHs (5.0-10.0). Notably, Aga4436 retained more than 90%, 80%, and 35% of its maximum activity after incubation at 30 °C, 40 °C, and 50 °C for 144 h, respectively, which exhibited an excellent thermostability in medium-high temperatures. Besides, Aga4436 displayed a remarkable tolerance to acid and alkaline environments, as it retained more than 70% of its maximum activity at a wide range of pHs from 3.0 to 10.0 after incubation in tested pHs for 60 min. These desirable properties of Aga4436 could make Aga4436 attractive in the food and nutraceutical industries.
