The development of targeted drug delivery systems has been a pivotal area in nanomedicine, addressing challenges like low drug loading capacity, uncontrolled release, and systemic toxicity. This study aims to develop and evaluate dual-functionalized mesoporous silica nanoparticles (MSN) for targeted delivery of celecoxib, enhancing drug loading, achieving controlled release, and reducing systemic toxicity through amine grafting and imidazolyl polyethyleneimine (PEI) gatekeepers. MSN were synthesized using the sol–gel method and functionalized with (3-aminopropyl) triethoxysilane (APTES) to create amine-grafted MSN (MSN-NH₂). Celecoxib was loaded into MSN-NH₂, followed by conjugation of imidazole-functionalized PEI (IP) gatekeepers synthesized via carbodiimide coupling. Characterization was conducted using Fourier-transform infrared spectroscopy (FTIR) and proton nuclear magnetic resonance (¹H-NMR). Drug loading capacity, entrapment efficiency, and in vitro drug release at pH 5.5 and 7.4 were evaluated. Cytotoxicity was assessed using the MTT assay on RAW 264.7 macrophages. The synthesized IP was confirmed by FTIR and ¹H-NMR. Amine-grafted MSN demonstrated a celecoxib loading capacity of 12.91 ± 2.02%, 2.1 times higher than non-functionalized MSN. In vitro release studies showed pH-responsive behavior with significantly higher celecoxib release from MSN-NH₂-celecoxib-IP at pH 5.5 compared to pH 7.4, achieving a 33% increase in release rate within 2 h. Cytotoxicity tests indicated significantly higher cell viability for IP-treated cells compared to PEI-treated cells, confirming reduced toxicity. The dual-functionalization of MSN with amine grafting and imidazolyl PEI gatekeepers enhances celecoxib loading and provides controlled pH-responsive drug release while reducing systemic toxicity. These findings highlight the potential of this advanced drug delivery system for targeted anti-inflammatory and anticancer therapies.

Endosomal entrapment significantly limits the efficacy of drug delivery systems. This study investigates sodium oleate-modified liposomes (SO-Lipo) as an innovative strategy to enhance endosomal escape and improve cytosolic delivery in 4T1 triple-negative breast cancer cells. We aimed to elucidate the mechanistic role of sodium oleate in promoting endosomal escape and compared the performance of SO-Lipo with unmodified liposomes (Unmodified-Lipo) and Aurein 1.2-modified liposomes (AUR-Lipo). Liposomes were prepared using the thin-film hydration method, resulting in Unmodified-Lipo, SO-Lipo, and AUR-Lipo formulations. The particle sizes were 102.2 ± 3.30 nm for Unmodified-Lipo, 109.6 ± 7.65 nm for SO-Lipo, and 151.9 ± 5.88 nm for AUR-Lipo, with polydispersity indices below 0.25, indicating uniform size distribution. Endosomal escape efficiency was evaluated through confocal microscopy by measuring the colocalization of labeled liposomes with lysosomal markers, quantified using Pearson’s correlation coefficient. Lipid mixing assays assessed the potential fusogenic effect, and molecular dynamics (MD) simulations explored the interactions of protonated sodium oleate (SO) with the endosomal membrane. Results demonstrated that SO-Lipo exhibited superior endosomal escape compared to Unmodified-Lipo, as evidenced by reduced colocalization with lysosomal markers, and achieved comparable efficacy to AUR-Lipo with lower cytotoxicity. Lipid mixing assays confirmed the potential fusogenic effect of SO with endosomal membrane models. MD simulations revealed that under acidic endosomal conditions, SO is protonated to oleic acid, which integrates into the membrane, enhancing fluidity and promoting fusion events essential for cytosolic release. SO-Lipo enhance endosomal escape through a fusogenic mechanism, facilitating cytosolic delivery with reduced cytotoxicity. This approach offers a safer and more effective option for targeted drug delivery applications.

Purpose: This paper introduces a complete in vitro investigation of cytotoxic and genotoxic effects of Phyllanthus niruri extract loaded chitosan nanoparticles (PNNP) on mouse Sertoli cell line (TM4), as well as their impact on spermatogenesis. Methods: Chitosan nanoparticles (ChNP) and PNNP were prepared using an ionic gelation process, while their cytotoxicity on TM4 cells was assessed using the Cell Counting Kit-8 (CCK-8) assay. Comet and fast halo assays were used to quantify single-strand DNA breaks in TM4 cells. To detect changes in cell morphology during apoptosis, nuclear staining with Hoechst 33342 was performed. An immunofluorescence assay was employed to examine the expression level of proteins connexin 43 and Claudin 11 in TM4 cells after exposure to PNNP concentration of 125 µg/mL. Results: The synthesized PNNP had a size of 170.6 nm, a polydispersity index of 0.269, a zeta potential of +37.8 mV, and a good entrapment efficiency of 71.0%. Encapsulation of Phyllanthus niruri into ChNP induced DNA damage in TM4 cells as determined by alkaline comet and fast halo assay (FHA). Additionally, it stimulated apoptosis, as determined by changes in cell morphology by Hoechst 33342 staining. There was significant down regulation of blood-testis barrier (BTB) proteins in TM4 cells after exposure to PNNP which could compromise the integrity of BTB and subsequently disrupt spermatogenesis process in male. Conclusion: Our investigation confirms the cytotoxic and genotoxic effects of PNNP in TM4 cells, which could lead to spermatogenesis disruption and male infertility.

The messenger RNA (mRNA) therapeutic platform has transitioned from a theoretical concept to a clinically validated, transformative modality in contemporary medicine. The unprecedented success of the COVID-19 vaccines, which accelerated global recognition of this technology, has catalyzed an intensive and broad exploration across numerous therapeutic domains. This review provides an integrated examination of the molecular and engineering innovations that have underpinned the platform's maturation. Collective advances, including refinements in cap analog chemistry, the meticulous optimization of untranslated regions and the poly(A) tail, precise tuning of codon usage, and strategic nucleoside modification, have synergistically enhanced mRNA stability, translational efficiency, and immune tolerance. Concurrently, the parallel evolution of lipid nanoparticles (LNPs) design, specifically the development of ionizable and biodegradable lipids, has established a reproducible and safe framework for highly efficient in vivo delivery. Furthermore, the review considers next-generation constructs, such as circular RNA (circRNA) and self-amplifying RNA (saRNA), which hold the distinct promise of extending expression durability and substantially reducing required dosing regimens. Emerging applications now extend beyond vaccination to oncology, rare genetic and metabolic diseases, and immune modulation. Despite these advances, major challenges persist in achieving extrahepatic delivery, improving endosomal escape, and enabling thermostable, scalable manufacturing. Continued integration of molecular engineering, data-driven design, and nanotechnology is expected to transform mRNA into a modular, programmable therapeutic platform. Collectively, these developments position mRNA therapeutics at the forefront of precision and regenerative medicine, defining a new trajectory for next-generation biologics.

The therapeutic benefits of Phyllanthus niruri (PN) are well-known, particularly in traditional medicine. Nanoemulsion technology has enhanced its bioavailability and efficacy, but potential nanotoxic effects, especially on male reproductive health, are not fully understood. This study examines the cytotoxic and genotoxic impacts of Phyllanthus niruri nanoemulsions (PNNE) of different sizes on TM4 Sertoli cells, focusing on their effects on the blood-testis barrier (BTB). PNNE of two distinct sizes, were prepared using spontaneous emulsification. The cytotoxicity of these formulations was assessed using a Cell Counting Kit-8 (CCK-8) assay, while genotoxicity was evaluated through an alkaline comet assay. Additionally, the expression of BTB proteins, claudin 11 and connexin 43, was examined via immunofluorescence analysis. PNNE were synthesized with droplet sizes of 16.9 ± 3.31 nm and 163.7 ± 8.53 nm. The smaller PNNE exhibited higher cytotoxicity (IC50 = 160.6 ± 8.3 μg/mL) compared to the larger PNNE (IC50 = 324.4 ± 12.5 μg/mL) and caused more significant DNA damage, as evidenced by the comet assay Both sizes led to a reduction in BTB protein expression, with the smaller nanoparticles causing more pronounced disruption. The study highlights the crucial role of nanoparticle size in determining the biological effects of PNNE on TM4 Sertoli cells. Smaller PNNE were found to be more detrimental to BTB integrity and cellular health, emphasizing the need for careful size optimization in the development of nanoemulsion-based therapies. These findings contribute to the understanding of nanotoxicity in the context of male reproductive health.

Background: Diabetic nephropathy (DN) is largely driven by transforming growth factor-β1 (TGF-β1)-mediated fibrosis. Clustered regularly interspaced short palindromic repeats (CRISPR)-associated protein 9 (Cas9) ribonucleoprotein (RNP) complexes offer precise gene disruption, yet effective non-viral delivery remains a challenge. This study developed cationic lipid-based hybrid nanostructured lipid carriers (NLCs) for intracellular delivery of TGFB1-targeting RNP as an early-stage platform for DN gene modulation. Methods: A single-guide RNA (sgRNA) targeting human TGFB1 was assembled with Cas9 protein (1:1 and 1:2 molar ratios). Hybrid NLCs comprising squalene, glyceryl trimyristate, and the cationic lipid 1,2-dioleoyl-3-trimethylammonium-propane (DOTAP) were formulated via optimized emulsification–sonication to achieve sub-100 nm particles. Physicochemical properties, including polydispersity index (PDI), were assessed via dynamic light scattering (DLS), while silencing efficacy in HEK293T cells was quantified using quantitative reverse transcription PCR (RT-qPCR) and enzyme-linked immunosorbent assay (ELISA). Results: Optimized NLCs achieved hydrodynamic diameters of 65–99 nm (PDI < 0.5) with successful RNP complexation. The 1:2 Cas9:sgRNA formulation produced the strongest gene-editing response, reducing TGFB1 mRNA by 67% (p < 0.01) compared with 39% for the 1:1 ratio. This translated to a significant reduction in TGF-β1 protein (p < 0.05) within 24 h. Conclusions: DOTAP-based hybrid NLCs enable efficient delivery of CRISPR–Cas9 RNP and achieve significant suppression of TGFB1 expression at both transcriptional and protein levels. These findings establish a promising non-viral platform for upstream modulation of profibrotic signaling in DN and support further evaluation in kidney-derived cells and in vivo renal models.