A Kidney-Targeted Nanoparticle to Augment Renal Lymphatic Density Decreases Blood Pressure in Hypertensive Mice

Chronic interstitial inflammation and renal infiltration of activated immune cells play an integral role in hypertension. Lymphatics regulate inflammation through clearance of immune cells and excess interstitial fluid. Previously, we demonstrated increasing renal lymphangiogenesis prevents hypertension in mice. We hypothesized that targeted nanoparticle delivery of vascular endothelial growth factor-C (VEGF-C) to the kidney would induce renal lymphangiogenesis, lowering blood pressure in hypertensive mice. A kidney-targeting nanoparticle was loaded with a VEGF receptor-3-specific form of VEGF-C and injected into mice with angiotensin II-induced hypertension or LNAME-induced hypertension every 3 days. Nanoparticle-treated mice exhibited increased renal lymphatic vessel density and width compared to hypertensive mice injected with VEGF-C alone. Nanoparticle-treated mice exhibited decreased systolic blood pressure, decreased pro-inflammatory renal immune cells, and increased urinary fractional excretion of sodium. Our findings demonstrate that pharmacologically expanding renal lymphatics decreases blood pressure and is associated with favorable alterations in renal immune cells and increased sodium excretion.

Suppression of fibrin(ogen)-driven pathologies through controlled knockdown by lipid nanoparticle delivery of siRNA

Fibrinogen plays a pathologic role in multiple diseases. It contributes to thrombosis and modifies inflammatory and immune responses, supported by studies in mice expressing fibrinogen variants with altered function or with a germline fibrinogen deficiency. However, therapeutic strategies to safely and effectively tailor plasma fibrinogen concentration are lacking. Here, we developed a strategy to tune fibrinogen expression by administering lipid nanoparticle (LNP)-encapsulated siRNA targeting the fibrinogen α chain (siFga). Three distinct LNP-siFga reagents reduced both hepatic Fga mRNA and fibrinogen levels in platelets and plasma, with plasma levels decreased to 42%, 16% and 4% of normal within one-week of administration. Using the most potent siFga, circulating fibrinogen was controllably decreased to 32%, 14%, and 5% of baseline with a 0.5, 1, and 2 mg/kg dose, respectively. Whole blood from mice treated with siFga formed clots with significantly decreased clot strength ex vivo, but siFga treatment did not compromise hemostasis following saphenous vein puncture or tail transection. In an endotoxemia model, siFga suppressed the acute phase response and decreased plasma fibrinogen, D-dimer, and proinflammatory cytokine levels. In a sterile peritonitis model, siFga restored normal macrophage migration in plasminogen-deficient mice. Finally, treatment of mice with siFga decreased the metastatic potential of tumour cells in a manner comparable to that observed in fibrinogen-deficient mice. The results indicate that siFga causes robust and controllable depletion of fibrinogen and provide the proof-of-concept that this strategy can modulate the pleiotropic effects of fibrinogen in relevant disease models.

Identifying target sites for placental therapeutics through the comparison of normal term pregnancies and intrauterine growth restricted proteomes

A variety of pathologies, including intrauterine growth restriction (IUGR), have been linked to placental insufficiencies as important causal factors, however, little has been done to develop therapeutics that may improve placental development, structure and function. The placenta offers the opportunity to manipulate the in-utero environment without directly intervening with the fetus, accessible from the maternal circulation, a vital but temporary organ, the placenta is no longer required after birth. Developing therapeutics must involve multiple aspects of targeting and safety to ensure no off-target impact on the pregnant person or fetus as well as enhance efficiency of delivery. In addition to our studies on nanoparticle delivery to the placenta [1] we are developing targeting strategies to allow peripheral delivery via the maternal circulation. In this study we have performed the isolation of the microvillous membrane from human placental syncytiotrophoblast (the targeting cell) and via proteomic analysis identified potential targeting moieties, we have then compared this to publicly available data from pregnancies complicated by fetal growth restriction to ensure that we do not identify targets which would be reduced in FGR, resulting in less efficient targeting.

A novel mechanism for the loss of mRNA activity in lipid nanoparticle delivery systems

Lipid nanoparticle (LNP)-formulated mRNA vaccines were rapidly developed and deployed in response to the SARS-CoV-2 pandemic. Due to the labile nature of mRNA, identifying impurities that could affect product stability and efficacy is crucial to the long-term use of nucleic-acid based medicines. Herein, reversed-phase ion pair high performance liquid chromatography (RP-IP HPLC) was used to identify a class of impurity formed through lipid:mRNA reactions; such reactions are typically undetectable by traditional mRNA purity analytical techniques. The identified modifications render the mRNA untranslatable, leading to loss of protein expression. Specifically, electrophilic impurities derived from the ionizable cationic lipid component are shown to be responsible. Mechanisms implicated in the formation of reactive species include oxidation and subsequent hydrolysis of the tertiary amine. It thus remains critical to ensure robust analytical methods and stringent manufacturing control to ensure mRNA stability and high activity in LNP delivery systems.

Nanotechnology applications in plant tissue culture and molecular genetics: A holistic approach

: Nanotechnology is one of the most important modern sciences that has integrated all sectors of science. Nanotechnology has been applied in the agricultural sector in the last ten years in pursuit of increasing agricultural production and ensuring food security. Plant biotechnology is an essential science that is concerned with plant production. The use of nanotechnology in plant biotechnology under controlled conditions has facilitated the understanding of important internal mechanisms of the plant biological system. The application of nanoparticles (NPs) in plant biotechnology has demonstrated an interesting impact on in vitro plant growth and development. This includes the positive effect of the NPs on micropropagation, callus induction, somatic embryogenesis, cell suspension culture, and plant disinfection. In addition, other biotechnology processes, including the genetic transformation of plants, plant conservation, and secondary metabolite production have improved by the use of NPs. Furthermore, nanotechnology is used to improve plant tolerance to different stress conditions that limit plant production. In this review article, we attempt to consolidate the achievements of nanotechnology and plant biotechnology and discuss advances in the applications of nanotechnology in plant biotechnology. It has been concluded that more research is needed to understand the mechanism of nanoparticle delivery and translocation in plants in order to avoid any future hazardous effects of nanomaterials. This will be key to the achievement of magnificent progress in plant nanobiotechnology.

Mimicking the Biology of Engineered Protein and mRNA Nanoparticle Delivery Using a Versatile Microfluidic Platform

To investigate the delivery of next-generation macromolecular drugs, such as engineered proteins and mRNA-containing nanoparticles, there is an increasing push towards the use of physiologically relevant disease models that incorporate human cells and do not face ethical dilemmas associated with animal use. Here, we illustrate the versatility and ease of use of a microfluidic platform for studying drug delivery using high-resolution microscopy in 3D. Using this microfluidic platform, we successfully demonstrate the specific targeting of carbonic anhydrase IX (CAIX) on cells overexpressing the protein in a tumor-mimicking chip system using affibodies, with CAIX-negative cells and non-binding affibodies as controls. Furthermore, we demonstrate this system’s feasibility for testing mRNA-containing biomaterials designed to regenerate bone defects. To this end, peptide- and lipid-based mRNA formulations were successfully mixed with colloidal gelatin in microfluidic devices, while translational activity was studied by the expression of a green fluorescent protein. This microfluidic platform enables the testing of mRNA delivery from colloidal biomaterials of relatively high densities, which represents a first important step towards a bone-on-a-chip platform. Collectively, by illustrating the ease of adaptation of our microfluidic platform towards use in distinct applications, we show that our microfluidic chip represents a powerful and flexible way to investigate drug delivery in 3D disease-mimicking culture systems that recapitulate key parameters associated with in vivo drug application.