Dedifferentiated Fat Cells as a Novel Source for Cell Therapy to Target Neonatal Hypoxic-Ischemic Encephalopathy

Abstract

Neonatal hypoxic-ischemic (HI) encephalopathy (HIE) remains a major cause of mortality and persistent neurological disabilities in affected individuals. At present, hypothermia is considered to be the only applicable treatment option, although growing evidence suggests that cell-based therapy might achieve better outcomes. Dedifferentiated fat (DFAT) cells are derived from mature adipocytes via a dedifferentiation strategy called ceiling culture. Their abundance and ready availability might make them an ideal therapeutic tool for the treatment of HIE. In the present study, we aimed to determine whether the outcome of HIE can be improved by DFAT cell treatment. HI injury was achieved by ligating the left common carotid artery in 7-day-old rat pups, followed by 1-h exposure to 8% O₂. Subsequently, the severity of damage was assessed by diffusion-weighted magnetic resonance imaging to assign animals to equivalent groups. 24 h after hypoxia, DFAT cells were injected at 10⁵ cells/pup into the right external jugular vein. To evaluate brain damage in the acute phase, a group of animals was sacrificed 48 h after the insult, and paraffin sections of the brain were stained to assess several acute injury markers. In the chronic phase, the behavioral outcome was measured by performing a series of behavioral tests. From the 24th day of age, the sensorimotor function was examined by evaluating the initial forepaw placement on a cylinder wall and the latency to falling from a rotarod treadmill. The cognitive function was tested with the novel object recognition (NOR) test. In vitro conditioned medium (CM) prepared from cultured DFAT cells was added at various concentrations to neuronal cell cultures, which were then exposed to oxygen-glucose deprivation (OGD). The number of cells that stained positive for the apoptosis marker active caspase-3 decreased by 73 and 52% in the hippocampus and temporal cortex areas of the brain, respectively, in the DFAT-treated pups. Similarly, the numbers of ED-1-positive cells (activated microglia) decreased by 66 and 44%, respectively, in the same areas in the DFAT-treated group. The number of cells positive for the oxidative stress marker 4-hydroxyl-2-nonenal decreased by 68 and 50% in the hippocampus and the parietal cortex areas, respectively, in the DFAT-treated group. The HI insult led to a motor deficit according to the rotarod treadmill and cylinder test, where it significantly affected the vehicle group, whereas no difference was confirmed between the DFAT and sham groups. However, the NOR test indicated no significant differences between any of the groups. DFAT treatment did not reduce the infarct volume, which was confirmed immunohistochemically. According to in vitro experiments, the cell death rates in the DFAT-CM-treated cells were significantly lower than those in the controls when DFAT-CM was added 48 h prior to OGD. The treatment effect of adding DFAT-CM 24 h prior to OGD was also significant. Our results indicate that intravenous injection with DFAT cells i...

Intravenous Administration of Bone Marrow-Derived Mesenchymal Stem Cell, but not Adipose Tissue-Derived Stem Cell, Ameliorated the Neonatal Hypoxic-Ischemic Brain Injury by Changing Cerebral Inflammatory State in Rat

“…Herein, intravenous injection of BM-MSCs reduced apoptosis induced by HI in the CA3 area and dentate gyrus of the hippocampus, the most vulnerable areas to HI insult in premature brain ( 37 ). This finding corresponds with those we recently reported using mononuclear cells derived from human umbilical cord blood cells ( 7 ) and dedifferentiated fat cells ( 32 ). Furthermore, BM-MSCs have been shown to exert a therapeutic effect through different administration routes, including intracranial ( 38 , 39 ), intracardiac ( 40 ), and intranasal ( 38 ).…”

“…HI rat models were made according to the method of Rice et al ( 31 ) with minor modification as described in our previous reports ( 7 , 32 ). On postnatal day 7 (P7), Wistar/ST male and female rat pups were anesthetized with isoflurane and their left common carotid artery was double-ligated with 5-0 surgical silk and cut between the ligatures.…”

“…The sample size was decided to be 5 to 10 in each group based on our previous studies ( 7 , 32 ). Statistical analyses were performed using JMP11.0 software (SAS Institute, Cary, NC, USA).…”

Adipocyte biology: It is time to upgrade to a new model

“…Cell sorting may improve the purity of cultured DFAT populations and the reproducibility of results obtained from these cells, but given that DFAT cells have a cell surface marker expression profile highly similar to that of ASCs and bone marrow MSCs (bmMSCs), it may prove difficult to define a set of DFAT‐specific cell surface markers (reviewed in Shah et al, ). DFAT cells are, perhaps somewhat surprisingly, not currently being used as a cell culture model for adipocyte biology, although several recent studies have utilized these cells in experimental stem cell‐based therapies (Mikrogeorgiou et al, ; D. Suzuki et al, ).…”

“…DFAT cells are, perhaps somewhat surprisingly, not currently being used as a cell culture model for adipocyte biology, although several recent studies have utilized these cells in experimental stem cellbased therapies (Mikrogeorgiou et al, 2017;D. Suzuki et al, 2017).…”