The bioactive compounds that are produced by leeches combine traditional and modern treatment since the saliva of the animal contains proteins and peptides with anticoagulant, anti-inflammatory, antimicrobial, antioxidant, and regenerative properties. In this review, their biochemical profile, mechanisms and clinical uses are considered with a special focus on the fact that they are utilized to combine traditional practices with the modern developments in biomedical approaches. Proteomic and transcriptomic research has recently found more than 100 bioactive molecules, such as hirudin, calin, eglins, bdellins and destabilase, which are related to the blood-feeding process and therapeutic processes. These compounds control blood clotting, control inflammatory mediators, block microbes and enhance wound healing and the development of new blood vessels. In clinical practice, leech therapy is common in the reconstruction and microsurgical practice to reduce venous congestion and enhance graft success. They are also shown to be useful in wound healing, cardiovascular health, musculoskeletal conditions and regenerative medicine, as well as emerging drug delivery systems of recombinant proteins and nanocarriers. Some of the challenges involve biological variation, infection or bleeding risks and stringent regulations on purity and standardization. Biotechnology has improved through other developments such as recombinant protein production, high-throughput omics, and nanotechnology, which will help resolve these problems, making them safe and scalable for clinical use. Altogether, leech bioactives are the prime examples of the sophisticated pharmacology of nature, which have the potential of being used as therapeutic agents in the future. The recent approach and incorporation in personalized medicine and bioengineering models reflect the leech's capacity to address complicated illness and unmet healthcare requirements to reassert its significance in preventive medicine and recent biomedicine.
Uncontrolled hemorrhage and delayed wound healing represent critical challenges in clinical practice. To address these issues, we developed a bioinspired honeysuckle-loaded mesoporous silica nanoparticle (HS-MSN) system that integrates sustained drug release with rapid hemostasis and pro-healing functions. Inspired by the unique surface structure of waxberries, HS-MSN was synthesized via a facile adsorption method, exhibiting high surface area, hierarchical porosity, and efficient loading of bioactive honeysuckle extract. The nanocomposite demonstrated a sustained release profile lasting up to 72 h, significantly enhancing the durability and bioavailability of therapeutic components. In vitro studies showed that HS-MSN accelerated clotting initiation within 10 s in both normal and hemophilic blood models, outperforming its individual components (bare MSN or honeysuckle extract alone). The material also exhibited excellent biocompatibility, hemocompatibility, and efficient cellular uptake. Moreover, the sustained release of honeysuckle constituents potently scavenged reactive oxygen species and suppressed pyroptosis by inhibiting NLRP3 inflammasome activation and pro-inflammatory cytokine release. In multiple murine injury models (tail amputation, liver wound, and limb amputation), HS-MSN achieved rapid hemostasis, significantly reduced blood loss, and shortened clotting time. Most notably, in a hemorrhagic full-thickness wound model, HS-MSN treatment resulted in substantially accelerated wound closure, with (67.02 ± 2.56)% healing achieved within 7 days, enhanced collagen deposition, and improved re-epithelialization, significantly outperforming control groups. The combination of sustained release capability, rapid hemostasis, and potent healing promotion makes HS-MSN a promising multifunctional nanotherapeutic for managing acute hemorrhagic wounds and facilitating tissue regeneration in emergency and surgical settings.
Hemorrhage remains the principal cause of death on the battlefield. It is suggested that Tranexamic acid (TXA) can improve survival of severely-bleeding casualties. The intravenous approach is not always available in the pre-hospital setting. It was shown that for every 15 min delay, the efficiency of TXA decreases by 10 %. This study was designed to assess the pharmacokinetic, pharmacodynamic, and pre-clinical efficacy of a TXA autoinjector in uncontrolled hemorrhage in swine. Non-compressible hemorrhage was induced by laparoscopic partial liver resection. TXA was administered intramuscularly by autoinjector (n = 25) or intravenously (control, n = 5). Blood levels of TXA and dynamics of clot formation were determined. Euthanasia was performed ninety minutes after injury followed by a laparotomy for the measurement of free blood and clots in the abdomen. The TXA levels in the autoinjector group exceeded the effective therapeutic threshold within <5 min and remained above the 10 mg/L threshold throughout the experiment. Intra-abdominal blood volumes, hemodynamic parameters, and indices of clot formation were similar between autoinjector-delivered and intravenouslyadministered groups. Autoinjector-based TXA provides sustained, anti-fibrinolytic levels within 2-5 min of administration in a swine model of uncontrolled hemorrhage emphasizing its important.
Thrombus formation within blood vessels poses a serious threat to human health and is closely associated with various cardiovascular disorders. Therefore, developing novel anticoagulants with reduced bleeding risk holds significant clinical importance. Factor XIa (FXIa), a key enzyme in the intrinsic coagulation pathway, has emerged as an attractive target for safer anticoagulant therapy. In this study, a series of novel small-molecule FXIa inhibitors were designed based on the scaffold of Asundexian through a bioisosteric replacement strategy. Starting from compound F22, structural optimization at the P2' region was conducted by replacing the amide with non-classical heterocycles, aiming to improve inhibitory potency, selectivity, and metabolic stability. Among the synthesized analogs, compound FE12 exhibited potent FXIa inhibition (IC50 = 4.4 nM), high selectivity over PKal (SI = 60.3), and favorable metabolic stability (T1/2 = 38.6 min in HLMs). Consistent with the in vitro enzyme assay, FE12 significantly prolonged aPTT in a dose-dependent manner, comparable to Asundexian. Molecular docking indicated that FE12 retains key hydrogen-bonding and water-mediated interactions within the FXIa active site, whereas its conformation in PKal shifts and loses these interactions, explaining its potent FXIa inhibitory activity and weak PKal inhibition. In the FeCl3-induced rat thrombosis model, FE12 effectively inhibited thrombus formation comparable to Asundexian. In the mouse tail bleeding assay, FE12 did not cause a significant prolongation of bleeding time, indicating a minimal effect on hemostasis. Furthermore, acute toxicity evaluation demonstrated its good safety and tolerability. Overall, FE12 exhibits an excellent balance of potency, selectivity and safety, representing a promising lead compound for the development of small-molecule FXIa inhibitors.
Polymer-drug conjugates (PDCs) represent a remarkable advancement in modern medicine, leveraging the physicochemical properties of polymers to enhance the efficacy and safety of therapeutic agents. This comprehensive review explores the historical evolution, selection criteria, design, and synthesis of PDC, highlighting their transformative impact on drug delivery. The Introduction of biocompatible and biodegradable polymers has been a game-changer, allowing seamless integration with biological systems. We delve into the various polymer selection criteria, focusing on commonly used polymers such as Polyethylene glycol (PEG), Poly Lactic Glycolic Acid (PLGA), and chitosan, and the importance of biocompatibility and biodegradability. The design and synthesis of PDC involve sophisticated chemical and physical conjugation methods, emphasising cleavable and non-cleavable linkers and targeting moieties. We explore the drug-release mechanisms critical for achieving targeted and controlled drug delivery, including pH-sensitive, enzyme-triggered, and redox-sensitive mechanisms. The advantages of PDCs, including improved pharmacokinetics and pharmacodynamics, enhanced targeted delivery, and reduced toxicity, are discussed in detail. Despite their numerous benefits, PDC face challenges, such as stability issues, manufacturing scalability, and immunogenicity. This review addresses these limitations and suggests potential solutions. We also highlight recent advances and innovations in the field, including stimuli-responsive systems, nano PDC, and emerging trends in precision medicine. The applications of PDCs extend beyond cancer therapy, including antimicrobial drug delivery and treatments for cardiovascular and neurodegenerative diseases. Regulatory and commercial aspects are examined, with a focus on FDA-approved PDCs, challenges in clinical translation, and market trends. Future directions for PDCs include integrating Artificial intelligence into conjugate design, developing hybrid systems with other drug-delivery technologies, and advancing sustainable synthesis approaches.
Chemotherapy, the first approach in breast cancer management, is limited owing to systemic toxicity and drug resistance. For instance, 5-fluorouracil in recommended doses cause severe side effects, highlighting the urgent necessity of finding more effective and safer combinations. Hence, this study aims to develop biocompatible natural-based nanocarriers for the co-delivery of loratadine, an antihistaminic drug along with 5-fluorouracil in order to enhance the anticancer efficacy while reducing the required dose of 5-fluorouracil. In silico virtual screening was performed to examine the probable molecular interactions between loratadine or 5-fluorouracil, individually with two different polymers, chitosan and zein, to determine the most suitable carrier system. Zein exhibited superior binding affinity compared to chitosan. Nanoparticle optimization was conducted using a Box-Behnken design with zein, tannic acid, and either loratadine or 5-fluorouracil concentration as independent variables. The optimized formulations were characterized by dynamic light scattering, entrapment efficiency, morphology, in-vitro release, followed by cytotoxicity, apoptosis, and cell-cycle analyses in MCF-7 cells. The optimal formulation consisted of zein (50 mg), tannic acid (131.93 mg), and loratadine or 5-fluorouracil (5 mg). The optimized formulation of Loratadine loaded nanoparticles (NPs) showed a particle size of 197 nm, polydispersity index (PDI) of 0.153, zeta potential of -21.78 mV, and entrapment efficiency of 61.33%. Furthermore, the optimized 5-fluorouracil loaded nanoparticles exhibited a particle size of 231 nm, 0.170 for PDI, zeta potential of -24.01 mV, and EE of 74.91% for entrapment efficiency. The sustained drug release profile exhibited a controlled pattern over 24-48 h. Flow cytometry results showed that the mixed nanoparticles exhibited potent cytotoxicity equivalent to 5-fluorouracil loaded nanoparticles alone despite containing only half the 5-fluorouracil dose, confirming a potential synergistic effect. These findings confirmed the potential of drug-loaded nanoparticles as promising drug delivery systems for breast cancer management.
This study aimed to develop Mahonia bealei alkaloid (MA)-loaded nanoparticles (NPs) targeting the blood-brain barrier (BBB) for the treatment of depression. The MA@NPs were prepared via self-assembly using transferrin-modified chitosan-N, O-carboxymethyl chitosan (NOCMS-CS-Tf) as the carrier. Formulation optimization yielded uniform MA@NPs (∼192 nm, +29 mV) with a drug loading of ∼16 % and encapsulation efficiency of ∼80 %. Transmission electron microscopy (TEM) and scanning electron microscopy (SEM) revealed spherical MA@NPs with an average size of ∼55 nm under TEM, rough and porous surfaces, and good dispersibility. X-ray diffraction (XRD) analysis showed that the MA@NPs exhibited amorphous characteristics with reduced crystallinity after MA loading, while Fourier transform infrared spectroscopy (FTIR) confirmed successful MA encapsulation via hydrogen bonding and intermolecular interactions. The MA@NPs remained stable for over one month at 4 °C. In vitro release assays showed a sustained-release profile under conditions mimicking the depressive state. Consistently, in vivo fluorescence imaging indicated prolonged systemic retention, corroborating the sustained-release behavior of the MA@NPs. Cellular uptake experiments indicated enhanced BBB-targeting ability of MA@NPs especially under conditions of cellular stress or damage. Compared to free MA, the MA@NPs significantly alleviated lipopolysaccharide (LPS)-induced neuroinflammation in BV2 microglial cells and corticosterone (CORT)-induced neuronal damage in PC12 cells. In vivo, superior anti-neuroinflammatory effects compared to free MA were demonstrated in an LPS-induced mouse model of depression. Network pharmacology analysis, supported by qRT-PCR validation, revealed that both MA and MA@NPs activated the cAMP signaling pathway by upregulating the mRNA expression of protein kinase A (PKA), cAMP response element-binding protein (CREB), and brain-derived neurotrophic factor (BDNF). In conclusion, MA@NPs enhance the antidepressant efficacy of MA through sustained drug release, targeted brain accumulation, and modulation of neuroinflammation and neuroplasticity-related pathways.
The 36-amino acid peptide known as neuropeptide Y (NPY) is widely expressed in both the central and peripheral nerve systems and is essential for regulating energy balance, stress responses, cardiovascular function, and immunological regulation through Gprotein- coupled Y receptors (Y1, Y2, Y4, Y5). To explore the various functions of NPY in hematological and cardiovascular disorders and investigate potential therapeutic approaches that target NPY signaling networks. A comprehensive literature analysis focused on NPY-mediated mechanisms in cardiovascular diseases (CVDs) and hematological disorders. The review highlights pharmacological modulators, including synthetic analogs, receptor-specific agents, enzyme inhibitors, and natural substances. NPY dysregulation promotes vasoconstriction and inflammation, particularly through Y1 receptor activation, contributing to diseases such as atherosclerosis, heart failure, and hypertension. In hematological disorders, NPY influences hematopoiesis, immune cell activity, and angiogenesis, affecting conditions such as thrombosis and leukemia. Therapeutic approaches include receptor-specific agonists and antagonists (e.g., [Leu31, Pro34]NPY, BAY 53-6206), enzyme inhibitors (DPP4, NEP), and natural substances (flavonoids, polyphenols, saponins). Although therapeutic resistance remains a challenge, glucocorticoids also affect NPY expression. NPY acts as a crucial modulator in hematological and cardiovascular disorders. Understanding its receptor-specific functions enables the development of targeted therapeutic strategies. Natural substances provide promising adjuncts for modulating NPY activity, supporting integrated approaches for treating NPY-related disorders.
Cardiovascular and cerebrovascular diseases (CCVDs) have become prominent global health threats, presenting substantial challenges due to their intricate pathological mechanisms and diverse clinical manifestations. Tanshinone IIA (TSA), an active compound derived from the traditional Chinese medicinal herb Salvia miltiorrhiza, exhibits notable therapeutic potential in these diseases due to its multifaceted mechanism of action. TSA protects the cardiovascular and cerebrovascular systems by inhibiting inflammation, reducing oxidative stress, preventing apoptosis and fibrosis, and modulating key signaling pathways, including toll‑like receptor 4/NF‑κB, PI3K/AKT and nuclear factor erythroid 2‑related factor 2/heme oxygenase‑1. Notably, considerable progress has been made in applying TSA to conditions such as atherosclerosis, myocardial infarction, heart failure and hypertension. The present review synthesizes current research on the molecular mechanisms of TSA in treating CCVDs and highlights innovations in nanodelivery systems (for example, rHDL, TPP‑TPGS/LPNs and CBSA‑PEG‑TSA‑NPs) that enhance its therapeutic efficacy by improving solubility, prolonging its half‑life and enhancing targeting capabilities. These advancements not only establish a foundation for the broader clinical application of TSA in CCVDs but also offer valuable insights for the development of new therapeutic agents.
Acorus calamus L., popularly called vacha or sweet flag, is used in Ayurveda, Unani, Siddha, Chinese and other traditional medicines to treat a wide range of illnesses, including neurological, gastrointestinal, respiratory, metabolic, kidney, and liver disorders. This study aims to investigate the vasorelaxation potential of β-asarone (BA), a key biomarker of the A. calamus, as well as BA-enriched essential oil (ACRO), supercritical CO2 extract (ACRE), and BA-free fraction of A. calamus. The vasoreactivity of BA, ACRO, ACRE, and the BA-free fraction of A. calamus was evaluated in an ex vivo system with isolated superior mesenteric atrial rings, including elucidating the mode of action. In vitro toxicity in rat smooth muscle cells and in vivo oral toxicity in Swiss albino mice were evaluated to determine the safety profile of the test extract and the molecule. BA, ACRO and ACRE exhibited promising vasorelaxation in isolated rat mesenteric arteries (1-30 μg/ml), whereas the BA-free fraction exhibited a negligible vasorelaxation response. BA-induced concentration-dependent relaxation responses were studied in rat mesenteric arteries, and it was found to be highly sensitive in modulating calcium channel function in vascular smooth muscle cells, suggesting L-type VDCC as a major putative target in vasorelaxation response. Further, BA, ACRO and ACRE were evaluated for cytotoxicity in vascular smooth muscle cells. In vitro toxicity of BA suggested its safety up to 30 μM in the MTT assay. However, ACRO and ACRE showed cytotoxicity and inhibition of cell proliferation beyond 30 μg/ml when incubated for 12h. In continuation, the results of in vivo toxicity showed that ACRO and ACRE exhibited potential toxicity and produced complete mortality with ACRO and partial mortality with ACRE within the observational period of seven days, although morbidity and observational changes were evident in all the treatment groups when given from the range of 300 mg/kg up to 2000 mg/kg. This data suggests a classic case of efficacy-toxicity paradox in A.calamus, suggesting further study to retain efficacy and enhance safety.
Foeniculum vulgare was investigated for the comparative phytochemical profiles and bioactivities of its aqueous extracts (decoction, E1; Soxhlet, E2) and essential oil (EO), together with in silico exploration of major constituents. Phenolics, flavonoids, and tannins were quantified spectrophotometrically, individual metabolites profiled by HPLC/UV-ESI-MS, and volatile components by GC-MS. Antioxidant capacity (DPPH, FRAP, TAC), antimicrobial activity (MIC/MBC or MFC against 29 bacteria and 8 fungi), and anticoagulant effects (prothrombin time [PT] and activated partial thromboplastin time [aPTT]) were assessed. Molecular docking targeted β-lactamases (OXA-10), thrombin, and myeloperoxidase. The decoction (E1) showed the highest phenolic (56.8 mg GAE/g) and flavonoid (22.8 mg QE/g) contents and superior radical-scavenging activity (DPPH IC50 = 228.8 µg/mL). The EO was dominated by (E)-anethole (79.7%) and fenchone (7.7%). The aqueous extract is rich in phenolic acids and flavonoids (quercetin-3-glucuronide 12.13%, chlorogenic acid 10.63%), typifying a caffeoylquinic acid-glucuronidated flavonol chemotype. Antimicrobial testing showed that the aqueous extract stopped Acinetobacter baumannii from growing (MIC = 600 µg/mL) and the EO stopped Candida parapsilosis from growing (MIC = 1200 µg/mL). E1 extended the aPTT to 73.7 s compared to 34.6 s in the control group, indicating disruption of the intrinsic coagulation pathway, whereas PT exhibited minimal alteration. Docking studies revealed that quercetin-3-glucuronide demonstrates significant binding affinity to β-lactamase OXA-10 (-6.4 kcal/mol) and thrombin (-7.4 kcal/mol), thereby substantiating its proposed roles in antibacterial and anticoagulant activities. In general, F. vulgare aqueous extracts and EO have antioxidant, antimicrobial, and anticoagulant properties that work well together. Phenolic glycosides and (E)-anethole are the main ingredients that make this happen. These results bolster translational opportunities for the development of cardiovascular, antimicrobial, and nutraceutical adjuncts, while emphasising the necessity for in vivo validation, bioavailability investigations, and long-term safety evaluations.
Carbonized Typhae Pollen (CTP) is a class of herbs that resolves blood stasis and stops bleeding. Blood stasis syndrome (BSS) is a pathological state associated with blood circulation disorders and vascular injury, which contributes to the development of cardiovascular diseases. The metabolic microenvironment of endothelial cells (ECs) plays a pivotal role in maintaining vascular homeostasis and is highly sensitive to pathological disturbances. CTP positively enhances the hemostatic phase via the vascular endothelial growth factor (VEGF)/phospholipase C gamma 1 (PLCγ1)/Ca2⁺/cyclooxygenase 2 (COX-2) pathway. Excessive hemostasis may increase the risk of vascular disease. Therefore, the dual pharmacological effect of CTP in achieving "stop bleeding without leaving stasis" implies the involvement of additional regulatory pathways that contribute to vascular protection. This study aims to elucidate the protective mechanisms of CTP against BSS-induced circulatory disorders and vascular injury. Models of BSS rats, zebrafish thrombosis model, and hypoxia-induced ECs model were used to evaluate vascular protection. Bioinformatics, metabolomic analyses, and molecular biology studies were integrated to investigate underlying mechanisms. Treatment with CTP promoted blood circulation, and ameliorated thrombosis in zebrafish. Furthermore, CTP markedly attenuated hypoxia and inflammation caused by blood stasis and facilitated vascular remodeling. Bioinformatics and metabolomic analyses suggested that VEGF and downstream arginine metabolism were key pathways. CTP enhanced vascular protection and promoted remodeling through the VEGF/PI3K/AKT signalling pathway, thereby facilitating vascular repair following injury. CTP mediated haemostasis through the VEGF/PLCγ1/Ca²⁺/COX-2 pathway while improving vascular function following haemostasis via the VEGF/PI3K/AKT pathway. These pathways function independently yet remain interconnected, jointly contributing to the vascular homeostasis.
Latexin (LXN), a naturally occurring low-molecular-weight protein, plays crucial roles in coagulation, inflammation, and thrombosis, making it highly relevant to hematological and cardiovascular research. In this study, we investigated the physiological function of LXN in coronary artery disease (CAD), with a particular focus on its heparin-dependent regulation of the coagulation process. Clinical evaluations showed that plasma LXN concentrations are significantly varied among patients with chronic coronary syndrome (CCS) versus acute coronary syndrome (ACS), which indicates its possible diagnostic and prognostic value. The animal experiments also showed that LXN deficiency significantly increases the time of bleeding and clotting, which suggests that LXN directly contributes to the formation of thrombus. Molecular docking and molecular dynamics simulations showed strong heparin-binding capacity (docking score: -5.923; binding energy: -78.512 kJ/mol), while circular dichroism spectroscopy confirmed that LXN-heparin complex formation induces conformational changes and enhances procoagulant activity. Together, these findings demonstrate that plasma LXN serves not only as a promising biomarker for CAD, particularly ACS, but also that LXN can competitively bind to heparin, reducing its interaction with AT-III and thereby diminishing heparin's anticoagulant effect, acting as a physiological regulator of the coagulation process. LXN, therefore, represents a potential new molecular target for early diagnosis and therapeutic intervention in CAD.
Multiple synthetic high-density lipoproteins (sHDLs) have been developed and extensively evaluated in preclinical and clinical trials, with their functionality potentially linked to specific lipid compositions. This study investigates how lipid composition influences sHDL interactions with platelets. We synthesized sHDL particles using ApoA1 mimetic peptide 22A complexed with various lipids (DMPC, POPC, DSPC, DPPC, and SM) differing in chain lengths, saturation levels, and transition temperatures. DMPC sHDL demonstrated superior inhibition of platelet aggregation across multiple agonist concentrations, while POPC sHDL showed limited efficacy only at lower thrombin concentrations. Interestingly, all formulations exhibited similar cholesterol removal abilities, and POPC sHDL demonstrated the highest platelet association despite its inferior antiplatelet effects. Mechanistic investigation revealed the involvement of phospholipase A2 (PLA2) enzymes in DMPC sHDL's potent antiplatelet effects. Inhibition of cytosolic PLA2 (cPLA2) and lipoprotein-associated PLA2 (Lp-PLA2) significantly reduced DMPC sHDL's antiplatelet activity. We demonstrated PLA2-mediated hydrolysis of DMPC sHDL, resulting in bioactive lipid metabolites, lysophosphatidylcholine (LPC) 14:0 and myristic acid, both in vitro and in vivo. These metabolites directly inhibited platelet aggregation, integrin activation, and α-granule secretion in a dose-dependent manner, with significantly greater potency than metabolites derived from other phospholipids. Our findings elucidate a novel mechanism by which sHDL's lipid composition influences its antiplatelet properties through the generation of bioactive lipid metabolites, offering insights for developing targeted cardiovascular therapies.
In a continued search for novel plant-based therapeutics with multi-target pharmacological potential, the medicinal plant Dischidia bengalensis (Apocynaceae) was investigated for the first time for its anti-inflammatory, analgesic, and thrombolytic properties, addressing critical therapeutic areas such as rheumatoid arthritis, acute pain, and thrombosis. The methanolic extract and solvent fractions (dichloromethane, n-hexane, and ethyl acetate) were evaluated through integrated in vivo, in vitro, and in silico approaches. Phytochemical screening and GC-MS profiling revealed a diverse array of bioactive constituents, including fatty acids, terpenoids, and phenolic derivatives, many of which are reported to exhibit pharmacological activities. In vivo assays demonstrated that the methanolic extract (400 mg/kg) markedly suppressed carrageenan-induced paw edema (92.31% inhibition) from the 2nd to 4th hour (p  <  0.05, p  <  0.01), while the n-hexane fraction produced the most pronounced analgesic response in both writhing and tail-immersion models (p  <  0.001). Furthermore, the methanolic extract displayed significant thrombolytic activity (33.38  ±  4.27% at 20 mg/mL, p < 0.001) in human blood clot lysis, suggesting potential application in cardiovascular disorders. The scientific novelty of this study was further underscored by in silico molecular docking, ADME/T, and PASS prediction studies. Key bioactive compounds, identified by GC-MS, showed strong binding affinities and promising drug-like properties against pivotal human targets such as TNF-α (PDB: 2AZ5), COX-2 (PDB: 6COX), and tissue plasminogen activator. These findings conclusively establish D. Bengalensis as a promising and novel source of lead compounds for the development of novel therapeutics against inflammatory, pain-related, and cardiovascular disorders.
Alzheimer's disease (AD) is a neurological illness that progresses over time and has limited therapeutic options. This is mostly because the blood-brain barrier prevents drugs from reaching the brain efficiently. This study sought to repurpose olmesartan (OLM), an angiotensin II receptor blocker exhibiting promising neuroprotective properties, by improving its brain-targeted delivery via intranasally administered bilosomes with chitosan shell. Chitosan coated bilosomes containing olmesartan (OLM-CTS-BLS) were formulated using a Box-Behnken design to optimize key variables (lecithin, sodium deoxycholate, and chitosan concentration) for achieving minimum diameter together with maximum entrapment efficiency and zeta potential. The optimized formulation demonstrated nanosized vesicles (184.89±6.11 nm), positive surface charge (+34.07±1.72 mV), and high encapsulation (87.36%±1.07). In vitro release showed sustained OLM delivery over 12 hours. Nasal diffusion studies conducted on sheep nasal mucosa demonstrated a significant increase in flux and permeability for OLM-CTS-BLS compared to OLM suspension. Pharmacokinetic studies in rats confirmed significantly improved brain bioavailability and a 1.4 fold increase in brain/plasma AUC ratio for OLM-CTS-BLS versus conventional routes. Behavioral assessments (Y-maze, Morris Water Maze) in lipopolysaccharide-induced AD rats revealed superior cognitive improvement in OLM-CTS-BLS-treated animals. Biochemical assays showed reduced acetylcholinesterase activity and lipid peroxidation. Histopathological and immunohistochemical evaluations demonstrated reduced amyloid-β deposition and preserved neuronal architecture without nasal toxicity. Collectively, these findings underscore the potential of intranasal OLM-CTS-BLS as a safe and effective drug delivery system for repurposing OLM in AD therapy by enhancing its brain delivery and therapeutic efficacy.
Salvia miltiorrhiza Bunge has been used traditionally for cardiovascular disorders, but its specific roles in stem cell cardiac differentiation remain unclear. In this study, we examined whether Salvia miltiorrhiza Bunge (SM) promotes cardiomyocyte differentiation from mouse embryonic stem cells (mESCs) and defined its underlying mechanism. To dynamically monitor cardiac differentiation, we established a Tnnt2-H2B-mCherry reporter mESC line that retained normal pluripotency and differentiation capacity. Using an embryoid body-based differentiation system, we found that SM exerted a distinct temporal effect on lineage progression: treatment during the early differentiation window inhibited pluripotency maintenance, proliferation, and mesodermal development, whereas administration during the cardiac precursor stage markedly enhanced cardiomyocyte formation, as indicated by increased beating embryoid bodies and upregulation of Isl1, Nkx2.5, Tnnt2, Myh6, and Myl7. Mechanistically, transcriptomic and protein analyses showed that SM suppressed canonical Wnt/β-catenin signaling, including downregulation of Dvl2, β-catenin, Axin2, c-Myc, and Cyclin D1, while Wnt activation WAY262611 partially reversed these effects. Further compound screening identified tanshinone IIA (Tan IIA) as the principal active constituent of SM, which largely recapitulated the pro-cardiogenic and Wnt-inhibitory effects of the crude extract. Together, these findings identify SM and Tan IIA as stage-dependent regulators of mESC fate and support their potential utility in natural product-based strategies for improving stem cell-derived cardiomyocyte generation.
Radiation-induced skin injury (RSI) is a critical complication of radiotherapy, affecting over 95% of patients and causing severe morbidity with treatment interruptions. Clinical therapies fail to address its multifactorial pathogenesis, including vascular dysfunction, oxidative stress, and chronic inflammation. Current clinical chitosan-based wound care materials face challenges like rapid degradation and insufficient mechanical stability. To tackle these, we developed a self-assembled carboxymethyl chitosan-crosslinked collagen hydrogel (Col+CMC) using traceable type I collagen from shad fish scales. This hydrogel forms a stable, porous hydrogel at physiological conditions without external crosslinkers, integrating collagen's structural support with carboxymethyl chitosan's iron-chelating properties. In a murine RSI model, Col+CMC accelerated ulcer repair by promoting collagen remodeling and endothelial angiogenesis, while suppressing oxidative stress and ferroptosis through activation of the GPX4-SLC7A11 pathway. In vitro studies confirmed its robust free radical scavenging capacity and pro-angiogenic effects on endothelial cells. The injectable, self-setting design offers clinical advantages: no crosslinking agents simplify application, traceable aquaculture-derived collagen ensures regulatory compliance, and the composite architecture enables multi-targeted therapy against RSI's pathological drivers. This work presents a biomaterial platform that merges self-assembly functionality with mechanistic targeting of iron homeostasis, addressing critical gaps in RSI management and showcasing translational potential for radiation therapy complications.
Blood-brain barrier (BBB) dysfunction is a well-established pathological phenotype of ischemic stroke, and targeting BBB integrity has emerged as a promising therapeutic strategy. Danshen-Chuanxiong formula (DS-CX), an effective herbal combination against ischemic stroke, has demonstrated regulatory effects on the BBB at various stages of ischemic stroke. However, its specific BBB-protective components and underlying molecular mechanisms remain unclear. Recent advances in multicellular self-assembled BBB spheroids have shown distinct advantages in disease modeling and drug discovery, offering a novel approach to address these questions. To simulate ischemic stroke-induced BBB dysfunction, we developed an oxygen-glucose deprivation/reoxygenation (OGD/R)-induced BBB disruption model using multicellular spheroids. To identify the effective substances of DS-CX responsible for BBB protection, we conducted a multi-parametric evaluation to assess BBB permeability, tight junctions, cell viability, reactive oxygen species (ROS) levels, inflammatory markers, and apoptotic phenotypes using high-content imaging. Further immunofluorescence and transcription analyses were performed to elucidate the BBB-protective mechanisms of DS-CX and its active components. Similar to the overall effects of DS-CX on BBB protection, preliminary screening fortunately found that both protocatechuic acid, ferulic acid, and senkyunolide I significantly reduced OGD/R-induced leakage, and upregulated the protein and mRNA levels of ZO-1 and Claudin-5 in BBB spheroids. Further multi-phenotypic assessments manifested that DS-CX and its active compounds effectively improved cell survival, reduced ROS production, inhibited inflammation, and decreased apoptosis, compared to the damaged BBB spheroids without drug intervention. Molecular experiments showed that DS-CX and its active constituents not only rescued the abnormal protein levels of pivotal targets related to oxidative stress (HO-1), inflammation (MMP-9, TLR-4), and apoptosis (Caspase-3, Bax, Bcl-2) in OGD/R-treated BBB spheroids, but also normalized the dysregulated mRNA levels of vWF, HO-1, MMP-9, TLR-4, TNF-α, IL-6, IL-1β, and IL-18 caused by OGD/R stimulation. Collectively, the present work successfully identified protocatechuic acid, ferulic acid, and senkyunolide I as key BBB-protective components of DS-CX against ischemic stroke. These compounds likely exert their therapeutic effects through multi-target regulation of oxidative stress, inflammation, and apoptosis. Our findings provide a novel spheroid-based multi-parametric screening approach for discovering BBB-targeted therapies in ischemic stroke.
Epilepsy management remains a significant clinical challenge, as conventional antiseizure medications primarily mitigate symptoms without addressing the core pathological drivers, specifically the vicious cycle formed by neuroinflammation and oxidative stress. Furthermore, the therapeutic efficacy of potential neuroprotective agents is severely compromised by the blood‒brain barrier (BBB), poor stability, and insufficient accumulation at epileptic lesions. Therefore, engineering a BBB‒penetrating delivery strategy that simultaneously disrupts the vicious cycle of neuroinflammation and oxidative stress is critical for achieving disease‒modifying effects in epilepsy treatment. Here, we developed a biomimetic, brain‒targeting nanosystem (R‒tFNAs@PB) by anchoring resveratrol‒loaded tetrahedral framework nucleic acids (tFNAs) onto a Prussian blue (PB) core. This nanosystem effectively traversed the BBB and exhibited precise accumulation within hippocampal epileptic foci. Mechanistically, R‒tFNAs@PB acted as a dual‒function modulator. The PB core and resveratrol synergistically scavenged reactive oxygen species (ROS) and activated the SIRT3/SOD2 signaling pathway, thereby increasing the mitochondrial antioxidant capacity. This cascade effectively inhibited NLRP3 inflammasome activation and promoted the polarization of microglia from the proinflammatory M1 phenotype to the anti‒inflammatory M2 phenotype. In a mouse model of kainic acid‒induced epilepsy, the nanosystem significantly reduced neuronal damage, reduced the seizure frequency and severity, and ameliorated cognitive deficits. This study presents a novel nanotherapeutic strategy that integrates microenvironment remodeling with neural repair. By leveraging the specific brain‒targeting capability of tFNAs and the synergistic antioxidant properties of the core‒shell structure, R‒tFNAs@PB represent a promising approach for treating refractory epilepsy through the precise regulation of the oxidative stress‒neuroinflammation axis via the SIRT3/SOD2 pathway.