Gadolinium doping enhances the photoacoustic signal of synthetic melanin nanoparticles: a dual modality contrast agent for stem cell imaging (Conference Presentation)

2019 ◽  
Author(s):  
Jeanne Lemaster ◽  
Jesse V. Jokerst
Keyword(s):  
Stem Cell ◽  
Contrast Agent ◽  
Cell Imaging ◽  
Photoacoustic Signal ◽  
Dual Modality ◽  
Gadolinium Doping ◽  
Melanin Nanoparticles

scholarly journals Exosome-like silica nanoparticles: a novel ultrasound contrast agent for stem cell imaging

Nanoscale ◽  
2017 ◽  
Vol 9 (1) ◽  
pp. 402-411 ◽  
Author(s):  
Fang Chen ◽  
Ming Ma ◽  
Junxin Wang ◽  
Fang Wang ◽  
Shi-Xiong Chern ◽  
...  
Keyword(s):  
Stem Cell ◽  
Contrast Agent ◽  
Silica Nanoparticles ◽  
Cell Imaging ◽  
Ultrasound Contrast Agent ◽  
Ultrasound Contrast

scholarly journals Sol–Gel Synthesis and Electrospraying of Biodegradable (P2O5)55–(CaO)30–(Na2O)15 Glass Nanospheres as a Transient Contrast Agent for Ultrasound Stem Cell Imaging

ACS Nano ◽  
2015 ◽  
Vol 9 (2) ◽  
pp. 1868-1877 ◽  
Author(s):  
Farzad Foroutan ◽  
Jesse V. Jokerst ◽  
Sanjiv S. Gambhir ◽  
Ophir Vermesh ◽  
Hae-Won Kim ◽  
...  
Keyword(s):  
Stem Cell ◽  
Contrast Agent ◽  
Cell Imaging ◽  
Sol Gel ◽  
Sol Gel Synthesis

scholarly journals Efficient Correction of HAX1 Mutations in Primary HSPCs of Severe Congenital Neutropenia Patients Using CRISPR/CAS9 GENE-Editing

Blood ◽  
2020 ◽  
Vol 136 (Supplement 1) ◽  
pp. 22-22
Author(s):  
Malte U Ritter ◽  
Benjamin Secker ◽  
Masoud Nasri ◽  
Maksim Klimiankou ◽  
Benjamin Dannenmann ◽  
...  
Keyword(s):  
Oxidative Stress ◽  
Bone Marrow ◽  
Stem Cell ◽  
Protein Expression ◽  
Healthy Donor ◽  
Live Cell Imaging ◽  
Gene Editing ◽  
Cell Imaging ◽  
Severe Congenital Neutropenia ◽  
Congenital Neutropenia

Patients with the rare pre-leukemia bone marrow failure syndrome severe congenital neutropenia (CN) have markedly reduced numbers of neutrophils in peripheral blood (<500/μl), leading to frequent infections and requiring chronic granulocyte stimulating factor (G-CSF) treatment. Approximately 7 % of CN patients carry homozygous loss-of-function mutations in the HAX1 gene. 25 % of HAX1-CN patients develop MDS or AML. The only curative therapy for CN patients with overt MDS/AML is hematopoietic stem cell transplantation with its associated risks. A clinical need for gene therapy for CN patients is imminent. Here, we describe for the first time the application of CRISPR/Cas9 gene-editing in combination with recombinant adeno associated virus 6 (rAAV6)-based delivery of the template for homology-directed repair (HDR) for the mutated HAX1 gene in primary bone marrow mononuclear CD34+ cells (HSPCs) of HAX1-CN patients. We selected HAX1 mutation p.W44X as the most frequently described mutation in HAX1-CN. We established the delivery of the chemically modified sgRNA in combination with SpCas9 V3 in primary HSPCs using electroporation. The HDR template was generated by PCR from healthy donor HSPCs and cloned into pRC6 vector for the production of high titer rAAV6 (>12x1012 viral copies per ml). Our gene-editing protocol produced on average 79,7 % (± 8,62 %) of total editing (TE) in healthy donor HSPCs (n=6). When we transduced healthy donor HSPCs with rAAV6 containing the template at MOI 105 after electroporation with CRISPR/Cas9 RNP, we achieved 38,1 % (± 1,3 %) knock-in (KI) efficiency and 82,3 % (± 8,2 %) TE (n=2). We further applied this approach to primary HSPCs from 5 CN patients harboring the p.W44X HAX1 mutation. We achieved 84,4 % (± 4,2 %) TE and 65,8 % (± 7,12 %) KI. Too proof, that our editing reintroduced HAX1 protein expression, we performed Western Blot analysis of edited cells (n=2) and were able to detect relevant amounts of HAX1 protein. To assess the effect of HAX1 correction on the neutropenic phenotype in vitro, we performed a liquid culture differentiation assay of edited HSPCs to neutrophils. HSPCs from the same patients that were edited in the AAVS1 safe harbor were used as isogenic controls. In the AAVS1 locus the editing efficiency was 76,74 % (± 17,07 %) total indels. By morphological assessment of Wright-Giemsa stained cytospins of edited cells derived on day 14 of differentiation revealed significant (p = 0,005) increases of mature neutrophils for all five edited HAX1-CN patient samples, as compared to the respective controls. This phenotype correction was also observed in flow cytometry by a significant (p = 0,011) increase of mature CD34-CD45+ CD15+CD16+ neutrophils (n=5). To investigate if the HAX1 mutation correction and reinforced expression of HAX1 protein improved the sensitivity of HSPCs to oxidative stress as described by Klein et al. 2007, we performed live-cell imaging of caspase3/7 activation. Live-cell imaging revealed a substantial reduction of H2O2-induced apoptosis in corrected HAX1-CN patients derived HSPCs (n=3). Furthermore, the corrected differentiated cells were investigated for functional hallmarks of granulocytes. We could observe that HAX1 gene-edited HSPCs showed comparable chemotaxis, phagocytosis and no defects in ROS production to isogenic control edited cells. Taken together, we established a protocol for efficient selection-free correction of HAX1 p.W44X mutation in primary HSPCs using CRISPR/Cas9 and rAVV6 HDR repair templates. Our gene-editing reintroduced HAX1 protein expression in primary HSPCs from HAX1-CN patients. Neutrophils derived from corrected cells showed functional improvements in survival to oxidative stress and general neutrophil functions. We believe that these results are enticing to be investigated further for potential clinical translation as an autologous stem cell therapy for HAX1-CN patients. Disclosures No relevant conflicts of interest to declare.

Stem Cell Imaging

2014 ◽  
pp. 4331-4338
Author(s):  
Francis L. Martin
Keyword(s):  
Stem Cell ◽  
Cell Imaging

Hydrogen peroxide degradable conjugated polymer nanoparticles for fluorescence and photoacoustic bimodal imaging

2018 ◽  
Vol 54 (20) ◽  
pp. 2518-2521 ◽  
Author(s):  
Jie Liu ◽  
Shaowei Wang ◽  
Xiaolei Cai ◽  
Shiwei Zhou ◽  
Bin Liu
Keyword(s):  
Hydrogen Peroxide ◽  
Contrast Agent ◽  
Conjugated Polymer ◽  
Cellular Imaging ◽  
Polymer Nanoparticles ◽  
Dual Modality ◽  
Bimodal Imaging ◽  
Conjugated Polymer Nanoparticles

A hydrogen peroxide degradable fluorescence/photoacoustic dual-modality contrast agent is prepared via in situ Sonogashira polymerization for cellular imaging.

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