Investigation into the usefulness of cynomolgus monkeys with spontaneously elevated intraocular pressure as a model for glaucoma treatment research.

Many glaucoma treatments focus on lowering intraocular pressure (IOP), with novel drugs continuing to be developed. One widely used model involves raising IOP by applying a laser to the trabecular iris angle (TIA) of cynomolgus monkeys to damage the trabecular meshwork. This model, however, presents challenges such as varying IOP values, potential trabecular meshwork damage, and risk of animal distress. This study investigated whether animals with naturally high IOP (>25 mmHg) could be used to effectively evaluate IOP-lowering drugs, thereby possibly replacing laser-induced models. Relationships between TIA size, IOP, and pupil diameter were also examined. Three representative IOP-lowering drugs (latanoprost, timolol, ripasudil) were administered, followed by multiple IOP measurements and assessment of corneal thickness, TIA, and pupil diameter via anterior segment optical coherence tomography (AS-OCT). There was a positive correlation was noted between IOP and corneal thickness before instillation, and a negative correlation between IOP and TIA before instillation. Our findings suggest animals with naturally high IOP could be beneficial for glaucoma research and development as a viable replacement for the laser-induced model and that measuring TIA using AS-OCT along with IOP yields a more detailed evaluation.

Matrix hyaluronic acid and bilayer poly-hydroxyethyl methacrylate-hyaluronic acid films as potential ocular drug delivery platforms.

This study aimed to design hydrogel based films comprising hyaluronic acid (HA) to overcome limitations of currently used eye drops. Timolol-loaded crosslinked (X2) HA-based and bilayer (B2) (pHEMA/PVP-HA-based layers) films were designed and characterized. The films were transparent (UV, visual observation) with crosslinked (<80 %) films showing lower light transmittance than bilayer (>80 %) films. X2 showed significantly higher swelling capacity, tensile strength and elastic modulus (5491.6 %, 1539.8 Nmm-2, 1777.2 mPa) than B2 (1905.0 %, 170.0N mm-2, 67.3 mPa) respectively. However, X2 showed lower cumulative drug released and adhesive force (27.3 %, 6.2 N) than B2 (57.5 %, 8.6 N). UV sterilization did not significantly alter physical properties, while SEM and IR microscopy showed smooth surface morphology and homogeneous drug distribution. Timolol permeation (EpiCorneal™/porcine cornea) depended on the film matrix with erodible films showing similar permeation to commercial eyedrops. Drug permeation for porcine cornea (X2 = 549.0.2, B2 = 312.1 µgcm-2 h-1) was significantly faster than EpiCorneal™ (X2 = 55.2, B2 = 37.6 µgcm-2 h-1), but with a linear correlation between them. All the selected optimized films showed acceptable compatibility (MTT assay) with both HeLa cells and EpiCorneal™. In conclusion, crosslinked and bilayer HA based films showed ideal characteristics suitable for potential ocular drug delivery, though further work is required to further optimize these properties and confirm their efficacy including in vivo tests.

The Role of Osteogenic Effect and Vascular Function in Bone Health in Hypertensive Rats: A Study of Anti-hypertensive and Hemorheologic Drugs.

Vascular dysfunction contributes to the development of osteopenia in hypertensive patients, as decreased blood supply to bones results in tissue damage and dysfunction. The effect of anti-hypertensive medicines on bone mass in hypertensive individuals is inconclusive because of the varied mechanism of their action, and suggests that reducing blood pressure (BP) alone is insufficient to enhance bone mass in hypertension. Pentoxifylline (PTX), a hemorheological drug, improves blood flow by reducing blood viscosity and angiogenesis, also has an osteogenic effect. We hypothesized that improving vascular function is critical to increasing bone mass in hypertension. To test this, we screened various anti-hypertensive drugs for their in vitro osteogenic effect, from which timolol and hydralazine were selected. In adult female spontaneously hypertensive rats (SHRs), timolol and hydralazine did not improve vascular function and bone mass, but PTX improved both. In female SHR animals, PTX restored bone mass, strength and mineralization, up to the level of normotensive control rats. In addition, we observed lower blood vasculature in the femur of adult SHR animals, and PTX restored them. PTX also restored the bone vascular and angiogenesis parameters that had been impaired in OVX SHR compared to sham SHR. This study demonstrates the importance of vascular function in addition to increased bone mass for improving bone health as achieved by PTX without affecting BP, and suggests a promising treatment option for osteoporosis in hypertensive patients, particularly at-risk postmenopausal women.

Dendritic Oligoethylenimine Decorated Liposome with Augmented Corneal Retention and Permeation for Efficient Topical Delivery of Antiglaucoma Drugs.

The topically administered glaucoma medications usually encounter serious precorneal drug loss and low corneal penetration, leading to a low bioavailability. In addition, due to the complexity of glaucoma etiology, a single medication is often insufficient. In this work, we report a novel dendritic oligoethylenimine decorated liposome for codelivery of two antiglaucoma drugs, latanoprost and timolol. The liposome showed a uniform nanoscopic particle size, positive surface charge, and excellent dual-drug loading capacity. A prolonged precorneal retention is observed by using this liposomal delivery system. This liposomal delivery system presents increased cellular uptake and tight junctions opening capacity, contributing respectively to the transcellular and paracellular permeation, thereby enhancing the trans-corneal transportation. Following topical administration of one eye drop in brown Norway rats, the dual-drug-loaded liposome formulation resulted in a sustained and effective intraocular pressure reduction as long as 5 days, without inducing ocular inflammation, discomfort, and tissue damage.

Efficacy of timolol in the treatment of facial hemangioma and its effect on the proliferation and apoptosis of hemangioma stem cells.

To investigate the efficacy of timolol in the treatment of facial hemangioma and the effect on the proliferation and apoptosis of hemangioma stem cells, 60 cases of children with IHs admitted to our hospital between 2020 and 2021 were selected and divided into two groups. The grouping was according to the lottery method, with 30 cases in each group. In the observation group, 0.5% timolol maleate eye drops were applied topically, and in the control group, propranolol hydrochloride tablets were administered orally to observe the efficacy of hemangioma, changes in hemangioma stem cells and the incidence of adverse reactions in both groups. Results showed that combined with the four-level score and ultrasound results, the number of effective treatment cases in the observation group was 28, which was higher than that in the control group, (P<0.05). The total number of adverse reactions in the observation group was 2, with an incidence rate. Under the intervention conditions of timolol, the proliferation level of hemangioma stem cells was inhibited, and the apoptosis rate of hemangioma stem cells increased with the increase of culture time (P<0.05). Among them, the apoptosis rate of the timolol group was higher than that of the blank control group at the same time point (P<0.05), and the difference was most significant at 48h (P<0.001). In conclusion, Timolol can effectively treat facial hemangioma in children, inhibit the proliferation of hemangioma stem cells and promote their apoptosis, with good curative effect, short treatment time and no obvious adverse reactions and it is economical and easy to accept.

Development of mucoadhesive Timolol loaded chitosan-nanocomposite to treat glaucoma.

Timolol Maleate is an aqueous soluble ß-blocker antiglaucoma drug used to suppress intraocular pressure. Several commercially available ocular formulations are not effective in delivering to the target site due to their water-soluble property and low mucoadhesiveness. Hence, there is a requirement for a highly mucoadhesive drug-loaded nanocomposite to suppress intraocular pressure with enhanced bioavailability. Herein, we have prepared a mucoadhesive Timolol-loaded graphene quantum dot-chitosan-nanocomposite to treat glaucoma in response to lysozyme, secreted in the tear fluid. The as-prepared nanocomposite has been characterized through high resolution-transmission electron microscopic, X-ray photoelectron spectroscopic, X-ray diffraction, and Fourier transform infrared spectral studies. The nanocomposite showed 93.74 % encapsulation efficiency with a loading capacity of 7.73 %. Further, 89.26 %, 95.62 %, and 99.29 % of drug release were observed from the nanocomposite in the presence of 1, 1.5, and 2 mg/mL of lysozyme. The mucoadhesion property has been confirmed by the increment in the particle size, fluorescence spectral variations, and Fourier transform infrared spectroscopic studies in the presence of mucin nanoparticles of size 291 nm. Interestingly, mucoadhesion has been demonstrated by pointing to the quenching in the luminescence of mucin. Further, in vitro biocompatibility assay on human corneal epithelial cells showed ≥80 % cell viability. Hence, this study offers the utilization of naturally secreting enzymes for drug delivery applications instead of uncontrolled pH and temperature-triggered releases.

RNA-seq analysis of human trabecular endothelial cells after treatment with timolol maleate.

PURPOSE: Timolol maleate (timolol), a ß-receptor blocker, reduces intraocular pressure by decreasing aqueous humor production. Timolol reportedly also protects ganglion cells, decreases aqueous humor outflow facility, and destroys the extracellular matrix in the trabecular meshwork. In this study, we investigated the effects of timolol on cultured human trabecular endothelial cells purchased from ScienCell using next-generation sequencing. STUDY DESIGN: Experimental investigation. METHODS: Total ribonucleic acid (RNA) was extracted after 24 h. More than 100 million RNAs in control and timolol-treated group were sequenced using a next-generation sequencer. The expression of 55,778 RNAs was analyzed. RESULTS: A total of 2,105 genes were significantly upregulated and 2,125 genes were downregulated, after the addition of timolol. VGF nerve growth factor inducible (VGF) (388-fold) had the maximum increase in expression, followed by amphiregulin (333-fold), a member of the epidermal growth factor family. Moreover, the expression of extracellular matrix-degrading enzymes, matrix metalloproteinases (MMPs) 1, 2, 3, 10, 12, and 14, increased. CONCLUSION: Timolol exerts various effects on human trabecular endothelial cells. The increase in MMP expression may contribute to the decrease in the aqueous humor outflow facility.

In Vivo Study of the Effects of Propranolol, Timolol, and Minoxidil on Burn Wound Healing in Wistar Rats.

Propranolol, timolol, and minoxidil have all shown benefits in treatment of burn injury and other skin wounds. The study evaluated their effects on full-thickness thermal skin burns in a Wistar rat model. Performed on 50 female rats; two dorsal skin burns were created on each animal. On the next day, the rats were divided into 5 groups (n = 10); each has received a specific treatment daily for 14 days: group I-topical vehicle (control), group II-topical silver sulfadiazine (SSD), group III-oral propranolol (5.5 mg) associated with topical vehicle, group IV-topical timolol 1% cream, and group V-topical minoxidil 5% cream. Wound contraction rates, malondialdehyde (MDA), glutathione (GSH, GSSG), and catalase activity in skin and/or serum were evaluated, and histopathological analyses were performed. Propranolol did not show advantages in necrosis prevention and wound contraction and healing, and did not reduce oxidative stress. It impaired keratinocyte migration, and promoted ulceration, chronic inflammation, and fibrosis, yet reducing the necrotic zone. Timolol prevented necrosis and promoted contraction and healing, increased antioxidant capacity and promoted keratinocyte migration and neo capillarization in comparison to the other treatments. Minoxidil reduced necrosis and enhanced contraction, resulting in positive outcomes after 1 week of treatment regarding local antioxidant defense, keratinocyte migration, neo capillarization, chronic inflammation, and fibrosis rates. However, after 2 weeks, it resulted in contrasting outcomes. In conclusion, topical timolol promoted wound contraction and healing, reducing local oxidative stress and improving keratinocyte migration, bringing arguments for potential benefits in skin epithelization.

Timolol maleate, a ß blocker eye drop, improved edema in a retinal vein occlusion model.

Purpose: To investigate the therapeutic effects of eye drops, namely, timolol maleate, a ß-adrenergic receptor antagonist, and latanoprost, a prostaglandin F2α analog, on retinal edema in a murine retinal vein occlusion (RVO) model. Methods: An RVO model was established using laser-induced RVO in mice, which were administered timolol maleate and latanoprost eye drops several times after venous occlusion. Subsequently, the thickness of the inner nuclear layer (INL) and the expression levels of such genes as Vegf and Atf4, which are stress markers of the endoplasmic reticulum, were examined. Primary human cultured retinal microvascular endothelial cells (HRMECs) were treated with timolol under hypoxic conditions, after which the gene expression pattern was investigated. Importantly, an integrated stress response inhibitor (ISRIB) was used in the RVO model, he known ISRIB, which suppresses the expression of ATF4 in retinal edema. Results: Increased INL thickness was suppressed by timolol eye drops, as were the expressions of Vegf and Atf4, in the RVO model. However, latanoprost eye drops did not induce any change in INL thickness. In HRMECs, hypoxic stress and serum deprivation increased the Vegf and Atf4 expressions; in response, treatment with timolol suppressed the Vegf expression. Furthermore, the ISRIB decreased the Vegf expression pattern and edema formation, which are associated with RVO. Conclusions: These results indicate that timolol eye drops may be a potential option for RVO treatment.

Activation of ADRB2/PKA Signaling Pathway Facilitates Lipid Synthesis in Meibocytes, and Beta-Blocker Glaucoma Drug Impedes PKA-Induced Lipid Synthesis by Inhibiting ADRB2.

Meibomian gland dysfunction is one of the main causes of dry eye disease and has limited therapeutic options. In this study, we investigated the biological function of the beta 2-adrenergic receptor (ADRB2)/protein kinase A (PKA) pathway in lipid synthesis and its underlying mechanisms in human meibomian gland epithelial cells (HMGECs). HMGECs were cultured in differentiation media with or without forskolin (an activator of adenylate cyclase), salbutamol (an ADRB2 agonist), or timolol (an ADRB2 antagonist) for up to 4 days. The phosphorylation of the cAMP-response element-binding protein (CREB) and the expression of peroxisome proliferator activator receptor (PPAR)γ and sterol regulatory element-binding protein (SREBP)-1 were measured by immunoblotting and quantitative PCR. Lipid synthesis was examined by LipidTOX immunostaining, AdipoRed assay, and Oil Red O staining. PKA pathway activation enhanced PPARγ expression and lipid synthesis in differentiated HMGECs. When treated with agonists of ADBR2 (upstream of the PKA signaling system), PPARγ expression and lipid synthesis were enhanced in HMGECs. The ADRB2 antagonist timolol showed the opposite effect. The activation of the ADRB2/PKA signaling pathway enhances lipid synthesis in HMGECs. These results provide a potential mechanism and therapeutic target for meibomian gland dysfunction, particularly in cases induced by beta-blocker glaucoma drugs.

Timolol 0.5% ophthalmic solution influences cardiac function in healthy cats.

OBJECTIVES: The aim of this study was to ascertain the effect of a drop of timolol 0.5% ophthalmic solution on the systolic function of the left ventricle (LV) and left atrium (LA), and to confirm if timolol helped appraisal of diastolic function by reducing heart rate (HR) and separating the transmitral outflow waves from tissue Doppler imaging (TDI). METHODS: A total of 41 client-owned healthy cats underwent two echocardiograms 20 mins apart. The timolol group (33 cats) received a drop of timolol solution after the first examination. Standard and speckle-tracking echocardiography evaluated the LV and LA function of both groups at the two time points evaluated. RESULTS: Timolol reduced HR (19%), and fractional shortening from LV (20.3%) and LA (16.6%). Septal S' decreased by 51% (from 7.7 to 5.2 cm/s) and lateral S' dropped by 43.1% (7.3 to 5.1 cm/s). Most longitudinal techniques did not change after timolol, including the mitral annular plane systolic excursion from the interventricular annulus, tricuspid annular plane systolic excursion, LV longitudinal strain and LV tissue motion annular displacement. The isovolumic relaxation time increased by 15.2% (from 54 to 64.6 ms), with most cats presenting this variable above the reference (>60 ms). Timolol did not support diastolic assessment, enabling evaluation in only 2/11 cats when using lateral TDI and 1/9 cats using septal TDI. Regarding side effects, miosis occurred in 18 cats (54.5%). CONCLUSIONS AND RELEVANCE: Timolol reduced systolic function, decreasing standard echocardiographic variables. Regarding diastolic evaluation, although timolol decreased HR, it did not separate the mitral diastolic waves, as expected.

Beta adrenergic receptor antagonist can modify Pseudomonas aeruginosa biofilm formation in vitro: Implications for chronic wounds.

Non-healing wounds are a major medical challenge, affecting over 6.5 million people in the US alone, with associated healthcare costs of about $16 billion annually. They can result in prolonged hospitalizations, work loss, disability, poor quality of life, and in diabetic patients with foot ulcers, amputation of the affected limb in 25% of patients. Though chronic ulcers may arise from different underlying diseases, the unifying feature is chronic infection, driving persistent inflammation that prolongs the healing process. One of the most frequently cultured or genetically identified pathogens in skin wounds is Pseudomonas aeruginosa. This species avidly forms biofilms in the wound that impede bacterial eradication by the host's immune mechanisms and limit efficacy of systemic antibiotics. Thus, non-antibiotic approaches to limit the growth and biofilm formation of this wound pathogen would be an advance in the treatment of chronic wounds. Prior work has demonstrated that the growth of other microbial species can be modulated by catecholamine agonists and antagonists of the adrenergic receptors (ARs). Here, we demonstrate that not only can the growth of this common wound pathogen be modulated by catecholamines, but also that the beta-AR antagonists can significantly decrease their growth, and importantly, limit their ability to form biofilms. These findings suggest that beta adrenergic antagonists may have a therapeutic role in the treatment of chronic skin wounds.

Combination product of dermal matrix, preconditioned human mesenchymal stem cells and timolol promotes wound healing in the porcine wound model.

A combination product of human mesenchymal stem/stromal cells (MSCs) embedded in an extracellular matrix scaffold and preconditioned with hypoxia and the beta-adrenergic receptor antagonist, timolol, combined with sustained timolol application post implantation, has shown promising results for improving wound healing in a diabetic mouse model. In the present study, we extend those findings to the more translatable large animal porcine wound model and show that the combined treatment promotes wound reepithelialization in these excisional wounds by 40.2% and increases the CD31 immunostaining marker of angiogenesis compared with the matrix control, while maintaining an accumulated timolol plasma concentration below the clinically safe level of 0.3 ng/mL after the 15-day course of topical application. Human GAPDH was not elevated in the day 15 wounds treated with MSC-containing device relative to wounds treated with matrix alone, indicating that the xenografted human MSCs in the treatment do not persist in these immune-competent animals after 15 days. The work demonstrates the efficacy and safety of the combined treatment for improving healing in the clinically relevant porcine wound model.

Effect of Topically Applied 0.5% Apraclonidine Versus 0.5% Timolol Maleate on Intraocular Pressure of Healthy Horses.

This study aims to assess the effect of topical 0.5% apraclonidine on Intraocular pressure (IOP) in horses and compare the effects of timolol maleate 0.5% with 0.5% apraclonidine in the equine eye. Twenty healthy female thoroughbred horses were used. Horses were divided into two groups. Ten horses received single dose of 0.2 mL of 0.5% apraclonidine in one randomly selected eye and the contralateral eye received single dose of 0.2 mL of artificial tears. In the second group, 10 horses received single dose of 0.2 mL of 0.5% timolol maleate in one eye and the opposite eye received single dose of placebo (0.2 mL of artificial tears). Intraocular pressure was measured using rebound tonometer at the baseline and 30, 60, 120, 240, 360 minutes, and 24 hours after topical ophthalmic drops instillation. Any ocular side effects were recorded at each time point. Mean (SD) baseline IOPs of the treated and placebo eyes were 26.2(3.1) and 23.5(3.4) in apraclonidine group, and 25.7(2.6) and 23.2(3.3) in timolol group. In the apraclonidine group, significant reduction in the mean IOP started after 60 minutes (P= .005) and was still present after 24 hours (P < .001). In timolol group, IOP was reduced in the treated eyes, but this reduction was only significant in the treated eyes at T24h (P= .03). The highest reduction in IOP in timolol group was observed at T360 (21.0(2.2); 14.7%). Mean IOP was decreased prominently by apraclonidine compared to timolol in treated eyes. In conclusion, single dose of topical 0.5% apraclonidine reduced IOP significantly among normal horses in the present study. Further investigations are necessary for evaluating efficacy and safety of apraclonidine in horses.

In Situ Gelling Electrospun Ocular Films Sustain the Intraocular Pressure-Lowering Effect of Timolol Maleate: In Vitro, Ex Vivo, and Pharmacodynamic Assessment.

Most common intraocular pressure (IOP) reduction regimens for the management of glaucoma include the topical use of eye drops, a dosage form that is associated with short residence time at the site of action, increased dosing frequency, and reduced patient compliance. In situ gelling nanofiber films comprising poly(vinyl alcohol) and Poloxamer 407 were fabricated via electrospinning for the ocular delivery of timolol maleate (TM), aiming to sustain the IOP-lowering effect of the ß-blocker, compared to conventional eye drops. The electrospinning process was optimized, and the physicochemical properties of the developed formulations were thoroughly investigated. The fiber diameters of the drug-loaded films ranged between 123 and 145 nm and the drug content between 5.85 and 7.83% w/w. Total in vitro drug release from the ocular films was attained within 15 min following first-order kinetics, showing higher apparent permeability (Papp) values across porcine corneas compared to the drug's solution. The fabricated films did not induce any ocular irritation as evidenced by both the hen's egg test on chorioallantoic membrane and the in vivo Draize test. In vivo administration of the ocular films in rabbits induced a faster onset of action and a sustained IOP-lowering effect up to 24 h compared to TM solution, suggesting that the proposed ocular films are promising systems for the sustained topical delivery of TM.

The Effects of Topical Timolol and Latanoprost on Calculated Ocular Perfusion Pressure in Nonglaucomatous Volunteers.

Purpose: To characterize the effects of timolol and latanoprost on calculated ocular perfusion pressure (OPP) in a multicenter, prospective, crossover-design study. Methods: Nonglaucomatous volunteers were evaluated at baseline, after 1 week of timolol 0.5% dosed twice daily, and after 1 week of latanoprost 0.005% dosed nightly (randomized treatment order; 6-week washout period). Pneumatonometric intraocular pressure (IOP) and brachial blood pressure (BP) were evaluated at each visit. Using 3 commonly used equations, OPP was calculated based on IOP and BP. The OPPs at each visit were compared by using linear mixed-effects models. Results: This analysis includes 121 participants (242 eyes; 75% female, 87% White, mean age 55 years). Mean OPP (standard deviation) calculated with mean arterial pressure was 46.8 (8.1) mmHg at baseline, 48.5 (7.9) mmHg with timolol (P = 0.005), and 49.6 mmHg (8.2) with latanoprost (P < 0.001). When compared with baseline, OPP calculated with diastolic BP was significantly increased with both timolol (1.3 mmHg) and latanoprost (3.1 mmHg). The OPP calculated with systolic BP was increased with latanoprost (2.8 mmHg) but decreased with timolol (-1.3 mmHg). Timolol reduced systolic BP by 3.2 mmHg. Compared with timolol, latanoprost conferred greater increases in OPP calculated with both systolic and diastolic BP compared with baseline; however, the difference in treatment effects on OPP calculated with mean arterial pressure was not significantly different (P = 0.068). Conclusion: In this crossover study of nonglaucomatous volunteers, latanoprost increased OPP. However, timolol's benefit to OPP may be limited in part because it reduced systolic BP. Clinical Trial Registration number: NCT01677507.

Morphological changes in the trabecular meshwork and Schlemm's canal after treatment with topical intraocular pressure-lowering agents.

Glaucoma treatment is usually initiated with topical medication that lowers the intraocular pressure (IOP) by reducing the aqueous production, enhancing the aqueous outflow, or both. However, the effect of topical IOP-lowering medications on the microstructures of the aqueous outflow pathway are relatively unknown. In this retrospective, observational study, 56 treatment-naïve patients with primary open-angle glaucoma were enrolled. Images of the nasal and temporal corneoscleral limbus were obtained using anterior segment optical coherence tomography (AS-OCT). The conjunctival vessels and iris anatomy were used as landmarks to select the same limbal area scan, and the trabecular meshwork (TM) width, TM thickness, and Schlemm's canal (SC) area were measured before and after using the IOP-lowering agents for 3 months. Among the 56 patients enrolled, 33 patients used prostaglandin (PG) analogues, and 23 patients used dorzolamide/timolol fixed combination (DTFC). After 3 months of DTFC usage, the TM width, TM thickness, and SC area did not show significant changes in either the nasal or temporal sectors. Conversely, after prostaglandin analog usage, the TM thickness significantly increased, and the SC area significantly decreased (all P < 0.01). These findings warrant a deeper investigation into their relationship to aqueous outflow through the conventional and unconventional outflow pathways after treatment with PG analogues.

Lollipop-Inspired Multilayered Drug Delivery Hydrogel for Dual Effective, Long-Term, and NIR-Defined Glaucoma Treatment.

Glaucoma is an ophthalmic disease that is characterized by elevated intraocular pressure (IOP). Eye drops are the preferred choice to reduce IOP for the treatment of glaucoma. However, the bioavailability of eye drops is low (<5%). Their long-term frequent administration cannot ensure patient compliance, which is the main reason for treatment failure. Inspired by lollipop, herein, a multilayered sodium alginate-chitosan (SA-CS) hydrogel ball (HB) decorated by zinc oxide-modified biochar (ZnO-BC) is developed as a new drug delivery system. The multilayer structure encapsulate timolol maleate (TM) and levofloxacin inside the different layers to realize the sustained release of drugs, which can control ocular hypertension and prevent infection effectively. The results show that the release of TM can be sustained in vitro for longer than 2 weeks. Moreover, IOP is also effectively reduced in vivo. Meanwhile, the photothermal conversion activity of ZnO-BC can regulate drug release on demand after stimulation by near-infrared irradiation. More importantly, the designed HB also shows good biocompatibility and antibacterial properties in vitro and in vivo. In summary, ZnO-BC-SA-CS HB can effectively reduce IOP and is expected to replace the classical tedious eye drop strategy, having potential utilization value in the treatment of glaucoma.

Timolol induces necroptosis, apoptosis and senescence concentration-dependently in rabbit Limbal stem cells in vitro.

Limbal stem cells (LSCs) are crucial for corneal transparency and vision. Any damages to LSCs might lead to limbal stem cell deficiency resulting in corneal opacification and even blindness. Here, we investigated the cytotoxicity of timolol and its underlying mechanisms in rabbit LSCs (rLSCs) in vitro. High concentrations of 0.5% and 0.25% timolol induced necroptosis in rLSCs to upregulate receptor interacting protein kinase (RIPK)1, RIPK3, mixed lineage kinase domain-like (MLKL) and phosphorylated MLKL along with downregulation of caspase-8 and caspase-2 within 4 h. While, median concentrations of 0.125% to 0.0625% timolol induced apoptosis in the rLSCs within 28 h. The apoptotic mechanism in the median-concentration timolol-treated rLSCs is probably via extrinsic apoptosis pathway by activating caspase-2, caspase-8 and caspase-3 and intrinsic apoptosis pathway triggered by excessive generation of ROS and subsequent DNA damage to upregulate Bax and Bad, downregulate Bcl-2 and Bcl-xL, subsequently disrupt mitochondrial membrane potential, cytosolically translocate cytochrome c and apoptosis-inducing factor, and activate caspase-9. In addition, low concentration of 0.03125% timolol induced senescence in the rLSCs by elevating ROS level and increasing number of senescence associated ß-galactosidase positive cells at 28 h. Our findings reveal that timolol induces necroptosis, apoptosis and senescence concentration-dependently in rLSCs in vitro.