Dataset Information

Rapid bioprinting of conjunctival stem cell micro-constructs for subconjunctival ocular injection.

ABSTRACT: Ocular surface diseases including conjunctival disorders are multifactorial progressive conditions that can severely affect vision and quality of life. In recent years, stem cell therapies based on conjunctival stem cells (CjSCs) have become a potential solution for treating ocular surface diseases. However, neither an efficient culture of CjSCs nor the development of a minimally invasive ocular surface CjSC transplantation therapy has been reported. Here, we developed a robust in vitro expansion method for primary rabbit-derived CjSCs and applied digital light processing (DLP)-based bioprinting to produce CjSC-loaded hydrogel micro-constructs for injectable delivery. Expansion medium containing small molecule cocktail generated fast dividing and highly homogenous CjSCs for more than 10 passages in feeder-free culture. Bioprinted hydrogel micro-constructs with tunable mechanical properties enabled the 3D culture of CjSCs while supporting viability, stem cell phenotype, and differentiation potency into conjunctival goblet cells. These hydrogel micro-constructs were well-suited for scalable dynamic suspension culture of CjSCs and were successfully delivered to the bulbar conjunctival epithelium via minimally invasive subconjunctival injection. This work integrates novel cell culture strategies with bioprinting to develop a clinically relevant injectable-delivery approach for CjSCs towards the stem cell therapies for the treatment of ocular surface diseases.

SUBMITTER: Zhong Z

PROVIDER: S-EPMC7719077 | biostudies-literature | 2021 Jan

REPOSITORIES: biostudies-literature

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Rapid bioprinting of conjunctival stem cell micro-constructs for subconjunctival ocular injection.

Zhong Zheng Z Deng Xiaoqian X Wang Pengrui P Yu Claire C Kiratitanaporn Wisarut W Wu Xiaokang X Schimelman Jacob J Tang Min M Balayan Alis A Yao Emmie E Tian Jing J Chen Luwen L Zhang Kang K Chen Shaochen S

Biomaterials 20201023

Ocular surface diseases including conjunctival disorders are multifactorial progressive conditions that can severely affect vision and quality of life. In recent years, stem cell therapies based on conjunctival stem cells (CjSCs) have become a potential solution for treating ocular surface diseases. However, neither an efficient culture of CjSCs nor the development of a minimally invasive ocular surface CjSC transplantation therapy has been reported. Here, we developed a robust in vitro expansio ...[more]

PMID: 33129190

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Project description:BACKGROUND:Keratomycosis is a relatively common, sight threatening condition in horses, where treatment is often prolonged and costly. Subconjunctival (SCo) injections offer less resistance to drug diffusion than the topical route, resulting in better penetration to the ocular anterior segment. Voriconazole, a second generation triazole antifungal, is effective against common fungal organisms causing keratomycosis. If combined with a thermogel biomaterial, voriconazole can be easily injected in the SCo space to provide sustained drug release. The purpose of this study was to evaluate the drug concentrations in the anterior segment and clinical effects after SCo injections of voriconazole-containing thermogel: poly (DL-lactide-co-glycolide-b-ethylene glycol-b-DL-lactide-co-glycolide) (PLGA-PEG-PLGA) in healthy equine eyes. RESULTS:Voriconazole aqueous humor (AH) and tear concentrations were compared between 6 horses, receiving 1% voriconazole applied topically (0.2?mL, q4h) (Vori-Top) or 1.7% voriconazole-thermogel (0.3?mL) injected SCo (Vori-Gel). For the Vori-Gel group, voriconazole concentrations were measured in AH and tears at day 2 and then weekly for 23?days, and at day 2 only for the Vori-Top group. Ocular inflammation was assessed weekly (Vori-Gel) using the modified Hackett-McDonald scoring system. Ocular tissue concentrations of voriconazole following SCo 1.7% voriconazole-thermogel (0.3?mL) injections were evaluated post euthanasia in 6 additional horses at 3 different time points. Three horses received bilateral injections at 2?h (n?=?3, right eye (OD)) and 48?h (n?=?3, left eye (OS)) prior to euthanasia, and 3 horses were injected unilaterally (OS), 7?days prior to euthanasia. Voriconazole-thermogel was easily injected and well tolerated in all cases, with no major adverse effects. On day 2, drug concentrations in tears were higher in the Vori-Top, but not statistically different from Vori-Gel groups. For the Vori-Gel group, voriconazole was non-quantifiable in the AH at any time point. Total voriconazole concentrations in the cornea were above 0.5??g/g (the target minimum inhibitory concentration (MIC) for Aspergillus sp.) for up to 48?h; however, concentrations were below this MIC at 7?days post treatment. CONCLUSIONS:Voriconazole-thermogel was easily and safely administered to horses, and provided 48?h of sustained release of voriconazole into the cornea. This drug delivery system warrants further clinical evaluation.

Project description:Mesenchymal stem cells (MSCs) have unique and beneficial properties and are currently used to treat a broad variety of diseases. These properties include the potential for differentiation into other cell types, secretion of different trophic factors that promote a regenerative microenvironment, anti-inflammatory actions, selective migration to damaged tissues, and non-immunogenicity. MSCs are effective for the treatment of ocular surface diseases such as dry eye, corneal burns, and limbal stem cell deficiency (LSCD), both in experimental models and in humans. LSCD is a pathological condition in which damage occurs to the limbal epithelial stem cells, or their niche, that are responsible for the continuous regeneration of the corneal epithelium. If LSCD is extensive and/or severe, it usually causes corneal epithelial defects, ulceration, and conjunctival overgrowth of the cornea. These changes can result in neovascularization and corneal opacity, severe inflammation, pain, and visual loss. The effectiveness of MSCs to reduce corneal opacity, neovascularization, and inflammation has been widely studied in different experimental models of LSCD and in some clinical trials; however, the methodological disparity used in the different studies makes it hard to compare outcomes among them. In this regard, the MSC route of administration used to treat LSCD and other ocular surface diseases is an important factor. It should be efficient, minimally invasive, and safe. So far, intravenous and intraperitoneal injections, topical administration, and MSC transplantation using carrier substrata like amniotic membrane (AM), fibrin, or synthetic biopolymers have been the most commonly used administration routes in experimental models. However, systemic administration carries the risk of potential side effects and transplantation requires surgical procedures that could complicate the process. Alternatively, subconjunctival injection is a minimally invasive and straightforward technique frequently used in ophthalmology. It enables performance of local treatments using high cell doses. In this review, we provide an overview of the current status of MSC administration by subconjunctival injection, analyzing the convenience, safety, and efficacy for treatment of corneal failure due to LSCD in different experimental models. We also provide a summary of the clinical trials that have been completed, are in progress, or being planned.

Project description:BackgroundT helper 2 (Th2) cells are thought to play critical roles in allergic conjunctivitis (AC). They release inflammatory cytokines to promote an allergic response in AC. Due to individual heterogeneity and long-term chronic management, current therapies do not always effectively control AC. Mesenchymal stem cells (MSCs) have been shown to be effective in treating allergy-related disorders, but it is unclear how exactly the Th2-mediated allergic response is attenuated. This study aims to elucidate the therapeutic effect and mechanism of the human umbilical cord MSCs (hUCMSCs) in a mouse model of experimental AC (EAC).MethodsA mouse EAC model was established by inoculating short ragweed (SRW) pollen. After the SRW pollen challenge, the mice received a single subconjunctival or tail vein injection of 2 × 106 hUCMSCs, or subconjunctival injection of hUCMSCs conditioned medium (hUCMSC-CM), and dexamethasone eye drops was used as positive control; subsequent scratching behavior and clinical symptoms were assessed. Immunostaining and flow cytometry were carried out to show allergic reactions and the activation of CD4 + T cell subsets in the conjunctiva and cervical lymph nodes (CLNs). Gene expression was determined by RNA-seq and further verified by qRT-PCR and Western blot. Co-culture assays were performed to explore the regulatory role of hUCMSCs in the differentiation of CD4 + naive T cells (Th0) into Th2 cells.ResultsSubconjunctival administration of hUCMSCs resulted in fewer instances of scratching and lower inflammation scores in EAC mice compared to the tail vein delivery, hUCMSC-CM and control groups. Subconjunctival administration of hUCMSCs reduced the number of activated mast cells and infiltrated eosinophils in the conjunctiva, as well as decreased the number of Th2 cells in CLNs. After pretreatment with EAC mouse serum in vitro to mimic the in vivo milieu, hUCMSCs were able to inhibit the differentiation of Th0 into Th2 cells. Further evidence demonstrated that repression of Th2 cell differentiation by hUCMSCs is mediated by CRISPLD2 through downregulation of STAT6 phosphorylation. Additionally, hUMCSCs were able to promote the differentiation of Th0 cells into regulatory T cells in CLNs of EAC mice.ConclusionsSubconjunctival injection of hUCMSCs suppressed the Th2-allergic response and alleviated clinical symptoms. This study provides not only a potential therapeutic target for the treatment of AC but also other T cell-mediated diseases.

Dataset Information

Rapid bioprinting of conjunctival stem cell micro-constructs for subconjunctival ocular injection.

Publications

Rapid bioprinting of conjunctival stem cell micro-constructs for subconjunctival ocular injection.

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