Quantitative assessment of surface functionality effects on microglial uptake and retention of PAMAM dendrimers

Liaw, Kevin; Gӧk, Özgül; DeRidder, Louis; Kannan, Sujatha; Kannan, Rangaramanujam M.

doi:10.1007/s11051-018-4219-1

Cited by 17 publications

(17 citation statements)

References 65 publications

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“…This may explain the more rapid and greater uptake of D4-OH by microglia rather than by astrocytes and other glial cell species in the mixed glial cell cultures. The earliest D4-OH PAMAM dendrimers primary glial cell internalization at 30 min was comparable to data previously reported previously by our group in immortalized BV2 mouse microglial cells [ 16 , 29 ]. Similar to our previous studies, we find that by 3 h most of the microglial cells have taken up the dendrimer [ 16 , 21 , 27 ].…”

Section: Discussionsupporting

confidence: 89%

“…However, the mechanisms by which this uptake occurs and the differences between ‘activated’ and ‘quiescent’ glial cells remain unexplored. Although it has been reported that the intracellular internalization of PAMAM dendrimers occurs via endocytosis, further investigation to determine the mechanisms of increased uptake by activated glial cells is necessary [ 28 , 29 ]. Recent studies have shown that besides the three key factors (surface charge, molecular weight, and generation), PAMAM dendrimer internalization may also be dependent on the targeted cell type [ 20 , 30 ].…”

Section: Introductionmentioning

confidence: 99%

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Preferential and Increased Uptake of Hydroxyl-Terminated PAMAM Dendrimers by Activated Microglia in Rabbit Brain Mixed Glial Culture

et al. 2018

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Polyamidoamine (PAMAM) dendrimers are multifunctional nanoparticles with tunable physicochemical features, making them promising candidates for targeted drug delivery in the central nervous system (CNS). Systemically administered dendrimers have been shown to localize in activated glial cells, which mediate neuroinflammation in the CNS. These dendrimers delivered drugs specifically to activated microglia, producing significant neurological improvements in multiple brain injury models, including in a neonatal rabbit model of cerebral palsy. To gain further insight into the mechanism of dendrimer cell uptake, we utilized an in vitro model of primary glial cells isolated from newborn rabbits to assess the differences in hydroxyl-terminated generation 4 PAMAM dendrimer (D4-OH) uptake by activated and non-activated glial cells. We used fluorescently-labelled D4-OH (D-Cy5) as a tool for investigating the mechanism of dendrimer uptake. D4-OH PAMAM dendrimer uptake was determined by fluorescence quantification using confocal microscopy and flow cytometry. Our results indicate that although microglial cells in the mixed cell population demonstrate early uptake of dendrimers in this in vitro system, activated microglia take up more dendrimer compared to resting microglia. Astrocytes showed delayed and limited uptake. We also illustrated the differences in mechanism of uptake between resting and activated microglia using different pathway inhibitors. Both resting and activated microglia primarily employed endocytotic pathways, which are enhanced in activated microglial cells. Additionally, we demonstrated that hydroxyl terminated dendrimers are taken up by primary microglia using other mechanisms including pinocytosis, caveolae, and aquaporin channels for dendrimer uptake.

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Section: Discussionsupporting

confidence: 89%

Section: Introductionmentioning

confidence: 99%

Preferential and Increased Uptake of Hydroxyl-Terminated PAMAM Dendrimers by Activated Microglia in Rabbit Brain Mixed Glial Culture

et al. 2018

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“…TPP-D-Cy5 exhibits highly punctated signal corresponding to the mitochondria marker, indicating a high degree of colocalization, as denoted by the yellow color signifying the overlapping of the red dendrimer signal and green mitochondrial signal ( Figure 3 A ). In contrast, D-Cy5 exhibits the expected diffuse cytosolic signal previously shown in both in vitro and in vivo macrophages 45 , 50 , 55 . D-Cy5 treated cells do exhibit some mitochondria-dendrimer signal overlap, but this overlap appears to arise from the broadness of cytosolic dendrimer signal rather than specific interactions with mitochondria.…”

Section: Resultsmentioning

confidence: 54%

“…Notably, the mitochondrial colocalization exhibited by D-Cy5 of approximately 45% is much greater than has been previously reported for non-targeting nanoparticles, which typically exhibit mitochondrial colocalization levels of 10-20%, and is similar to the colocalization levels exhibited by mitochondria-targeting modified versions of these nanoparticles of 40-60%. 6 , 23 This observed effect may partially arise from the relatively high cytosolic content of the dendrimer compared to other nanoparticles of similar size, which broadly overlaps with mitochondrial signal nonspecifically 45 , 58 , 59 . This suggests that hydroxyl-terminated PAMAM dendrimers are superior vehicles for intracellular targeting, and that their inherent intracellular targeting capabilities can be significantly enhanced through modification with targeting moieties.…”

Section: Resultsmentioning

confidence: 99%

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Targeting Mitochondrial Dysfunction and Oxidative Stress in Activated Microglia using Dendrimer-Based Therapeutics

et al. 2018

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Mitochondrial oxidative stress is associated with many neurodegenerative diseases, such as traumatic brain injury (TBI). Targeted delivery of antioxidants to mitochondria has failed to translate into clinical success due to their nonspecific cellular localization, poor transport properties across multiple biological barriers, and associated side effects. These challenges, coupled with the complex function of the mitochondria, create the need for innovative delivery strategies.Methods: Neutral hydroxyl-terminated polyamidoamine (PAMAM) dendrimers have shown significant potential as nanocarriers in multiple brain injury models. N-acetyl cysteine (NAC) is a clinically used antioxidant and anti-inflammatory agent which has shown significant potency when delivered in a targeted manner. Here we present a mitochondrial targeting hydroxyl PAMAM dendrimer-drug construct (TPP-D-NAC) with triphenyl-phosphonium (TPP) for mitochondrial targeting and NAC for targeted delivery to mitochondria in injured glia. Co-localization and mitochondrial content of mitochondria-targeted and unmodified dendrimer were assessed in microglia and macrophages in vitro via immunohistochemistry and fluorescence quantification. Therapeutic improvements of TPP-D-NAC over dendrimer-NAC conjugate (D-NAC) and free NAC were evaluated in vitro in microglia under oxidative stress challenge. In vivo neuroinflammation targeting was confirmed in a rabbit model of TBI.Results: TPP-conjugated dendrimer co-localized significantly more with mitochondria than unmodified dendrimer without altering overall levels of cellular internalization. This targeting capability translated to significant improvements in the attenuation of oxidative stress by TPP-D-NAC compared to D-NAC and free NAC. Upon systemic administration in a rabbit TBI model, TPP-conjugated dendrimer co-localized specifically with mitochondria in activated microglia and macrophages in the white matter of the ipsilateral/injured hemisphere, confirming its BBB penetration and glial targeting capabilities.Conclusion: D-NAC has shown promising efficacy in many animal models of neurodegeneration, and this work provides evidence that modification for mitochondrial targeting can further enhance its therapeutic efficacy, particularly in diseases where oxidative stress-induced glial cell death plays a significant role in disease progression.

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Targeted systemic dendrimer delivery of CSF‐1R inhibitor to tumor‐associated macrophages improves outcomes in orthotopic glioblastoma

Liaw

Reddy

Sharma

et al. 2020

Bioengineering & Transla Med

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Glioblastoma is the most common and aggressive form of primary brain cancer, with median survival of 16–20 months and a 5‐year survival rates of <5%. Recent advances in immunotherapies have shown that addressing the tumor immune profile by targeting the colony‐stimulating factor 1 (CSF‐1) signaling pathway of tumor‐associated macrophages (TAMs) has the potential to improve glioblastoma therapy. However, such therapies have shown limited successes in clinical translation partially due to lack of specific cell targeting in solid tumors and systemic toxicity. In this study, we present a novel hydroxyl dendrimer‐mediated immunotherapy to deliver CSF‐1R inhibitor BLZ945 (D‐BLZ) from systemic administration selectively to TAMs in glioblastoma brain tumors to repolarize the tumor immune environment in a localized manner. We show that conjugation of BLZ945 to dendrimers enables sustained release in intracellular and intratumor conditions. We demonstrate that a single systemic dose of D‐BLZ targeted to TAMs decreases pro‐tumor expression in TAMs and promotes cytotoxic T cell infiltration, resulting in prolonged survival and ameliorated disease burden compared to free BLZ945. Our results demonstrate that dendrimer‐drug conjugates can facilitate specific, localized manipulation of tumor immune responses from systemic administration by delivering immunotherapies selectively to TAMs, thereby improving therapeutic efficacy while reducing off‐target effects.

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Quantitative assessment of surface functionality effects on microglial uptake and retention of PAMAM dendrimers

Cited by 17 publications

References 65 publications

Preferential and Increased Uptake of Hydroxyl-Terminated PAMAM Dendrimers by Activated Microglia in Rabbit Brain Mixed Glial Culture

Preferential and Increased Uptake of Hydroxyl-Terminated PAMAM Dendrimers by Activated Microglia in Rabbit Brain Mixed Glial Culture

Targeting Mitochondrial Dysfunction and Oxidative Stress in Activated Microglia using Dendrimer-Based Therapeutics

Targeted systemic dendrimer delivery of CSF‐1R inhibitor to tumor‐associated macrophages improves outcomes in orthotopic glioblastoma

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