Nonhematopoietic Umbilical Cord Blood Stem Cell Administration Improves Long-term Neurodevelopment After Periventricular-Intraventricular Hemorrhage in Neonatal Rats.

Periventricular-intraventricular hemorrhage (PIVH) is common in extremely low gestational age neonates (ELGAN) and leads to motor and behavioral impairments. Currently there is no effective treatment for PIVH. Whether human nonhematopoietic umbilical cord blood-derived stem cell (nh-UCBSC) administration reduces the severity of brain injury and improves long-term motor and behavioral function was tested in an ELGAN-equivalent neonatal rat model of PIVH. In a collagenase-induced unilateral PIVH on postnatal day (P) 2 model, rat pups received a single dose of nh-UCBSCs at a dose of 1 × 106 cells i.p. on P6 (PIVH + UCBSC group) or were left untreated (Untreated PIVH group). Motor deficit was determined using forelimb placement, edge-push, and elevated body swing tests at 2 months (N = 5-8). Behavior was evaluated using open field exploration and rearing tests at 4 months (N =10-12). Cavity volume and hemispheric volume loss on the PIVH side were determined at 7 months (N = 6-7). Outcomes were compared between the Untreated PIVH and PIVH + UCBSC groups and a Control group. Unilateral motor deficits were present in 60%-100% of rats in the Untreated PIVH group and 12.5% rats in the PIVH + UCBSC group (P = 0.02). Untreated PIVH group exhibited a higher number of quadrant crossings in open field exploration, indicating low emotionality and poor habituation, and had a cavitary lesion and hemispheric volume loss on the PIVH side. Performance in open field exploration correlated with cavity volume (r2 = 0.25; P < 0.05). Compared with the Untreated PIVH group, performance in open field exploration was better (P = 0.0025) and hemispheric volume loss was lower (19.9 ± 4.4% vs 6.1 ± 2.6%, P = 0.018) in the PIVH + UCBSC group. These results suggest that a single dose of nh-UCBSCs administered in the subacute period after PIVH reduces the severity of injury and improves neurodevelopment in neonatal rats.

ssDNA nanotubes for selective targeting of glioblastoma and delivery of doxorubicin for enhanced survival.

Effective treatment of glioblastoma remains a daunting challenge. One of the major hurdles in the development of therapeutics is their inability to cross the blood-brain tumor barrier (BBTB). Local delivery is an alternative approach that can still suffer from toxicity in the absence of target selectivity. Here, we show that nanotubes formed from self-assembly of ssDNA-amphiphiles are stable in serum and nucleases. After bilateral brain injections, nanotubes show preferential retention by tumors compared to normal brain and are taken up by glioblastoma cells through scavenger receptor binding and macropinocytosis. After intravenous injection, they cross the BBTB and internalize in glioblastoma cells. In a minimal residual disease model, local delivery of doxorubicin showed signs of toxicity in the spleen and liver. In contrast, delivery of doxorubicin by the nanotubes resulted in no systemic toxicity and enhanced mouse survival. Our results demonstrate that ssDNA nanotubes are a promising drug delivery vehicle to glioblastoma.

Conduction Cooling and Plasmonic Heating Dramatically Increase Droplet Vitrification Volumes for Cell Cryopreservation.

Droplet vitrification has emerged as a promising ice-free cryopreservation approach to provide a supply chain for off-the-shelf cell products in cell therapy and regenerative medicine applications. Translation of this approach requires the use of low concentration (i.e., low toxicity) permeable cryoprotectant agents (CPA) and high post cryopreservation viability (>90%), thereby demanding fast cooling and warming rates. Unfortunately, with traditional approaches using convective heat transfer, the droplet volumes that can be successfully vitrified and rewarmed are impractically small (i.e., 180 picoliter) for <2.5 m permeable CPA. Herein, a novel approach to achieve 90-95% viability in micro-liter size droplets with 2 m permeable CPA, is presented. Droplets with plasmonic gold nanorods (GNRs) are printed onto a cryogenic copper substrate for improved cooling rates via conduction, while plasmonic laser heating yields >400-fold improvement in warming rates over traditional convective approach. High viability cryopreservation is then demonstrated in a model cell line (human dermal fibroblasts) and an important regenerative medicine cell line (human umbilical cord blood stem cells). This approach opens a new paradigm for cryopreservation and rewarming of dramatically larger volume droplets at lower CPA concentration for cell therapy and other regenerative medicine applications.

Zika virus-based immunotherapy enhances long-term survival of rodents with brain tumors through upregulation of memory T-cells.

Zika virus (ZIKV) exhibits a tropism for brain tumor cells and has been used as an oncolytic virus to target brain tumors in mice with modest effects on extending median survival. Recent studies have highlighted the potential for combining virotherapy and immunotherapy to target cancer. We postulated that ZIKV could be used as an adjuvant to enhance the long-term survival of mice with malignant glioblastoma and generate memory T-cells capable of providing long-term immunity against cancer remission. To test this hypothesis mice bearing malignant intracranial GL261 tumors were subcutaneously vaccinated with irradiated GL261 cells previously infected with the ZIKV. Mice also received intracranial injections of live ZIKV, irradiation attenuated ZIKV, or irradiated GL261 cells previously infected with ZIKV. Long-term survivors were rechallenged with a second intracranial tumor to examine their immune response and look for the establishment of protective memory T-cells. Mice with subcutaneous vaccination plus intracranial irradiation attenuated ZIKV or intracranial irradiated GL261 cells previously infected with ZIKV exhibited the greatest extensions to overall survival. Flow cytometry analysis of immune cells within the brains of long-term surviving mice after tumor rechallenge revealed an increase in the number of T-cells, including CD4+ and tissue-resident effector/ effector memory CD4+ T-cells, in comparison to long-term survivors that were mock-rechallenged, and in comparison to naïve untreated mice challenged with intracranial gliomas. These results suggest that ZIKV can serve as an adjuvant to subcutaneous tumor vaccines that enhance long-term survival and generate protective tissue-resident memory CD4+ T-cells.

Interspecies Organogenesis for Human Transplantation.

Blastocyst complementation combined with gene editing is an emerging approach in the field of regenerative medicine that could potentially solve the worldwide problem of organ shortages for transplantation. In theory, blastocyst complementation can generate fully functional human organs or tissues, grown within genetically engineered livestock animals. Targeted deletion of a specific gene(s) using gene editing to cause deficiencies in organ development can open a niche for human stem cells to occupy, thus generating human tissues. Within this review, we will focus on the pancreas, liver, heart, kidney, lung, and skeletal muscle, as well as cells of the immune and nervous systems. Within each of these organ systems, we identify and discuss (i) the common causes of organ failure; (ii) the current state of regenerative therapies; and (iii) the candidate genes to knockout and enable specific exogenous organ development via the use of blastocyst complementation. We also highlight some of the current barriers limiting the success of blastocyst complementation.

Immunomodulation with Human Umbilical Cord Blood Stem Cells Ameliorates Ischemic Brain Injury - A Brain Transcriptome Profiling Analysis.

Our group previously demonstrated that administration of a CD34-negative fraction of human non- hematopoietic umbilical cord blood stem cells (UCBSC) 48 h after ischemic injury could reduce infarct volume by 50% as well as significantly ameliorate neurological deficits. In the present study, we explored possible mechanisms of action using next generation RNA sequencing to analyze the brain transcriptome profiles in rats with ischemic brain injury following UCBSC therapy. Two days after ischemic injury, rats were treated with UCBSC. Five days after administration, total brain mRNA was then extracted for RNAseq analysis using Illumina Hiseq 2000. We found 275 genes that were significantly differentially expressed after ischemic injury compared with control brains. Following UCBSC treatment, 220 of the 275 differentially expressed genes returned to normal levels. Detailed analysis of these altered transcripts revealed that the vast majority were associated with activation of the immune system following cerebral ischemia which were normalized following UCBSC therapy. Major alterations in gene expression profiles after ischemia include blood-brain-barrier breakdown, cytokine production, and immune cell infiltration. These results suggest that UCBSC protect the brain following ischemic injury by down regulating the aberrant activation of innate and adaptive immune responses.