Compounding engineered mesenchymal stem cell-derived exosomes: A potential rescue strategy for retinal degeneration.

The prevalence of retinal degenerative diseases, including age-related macular degeneration and retinitis pigmentosa, has been increasing globally and is linked to the aging population and improved life expectancy. These diseases are characterized by chronic, progressive neuronal damage or depletion of the photoreceptor cells in the retina, and limited effective treatment options are currently available. Mesenchymal stem cell-derived exosomes (MSC-EXOs) containing cytokines, growth factors, lipids, mRNA, and miRNA, which act as mediators of intercellular communication transferring bioactive molecules to recipient cells, offer an appealing, non-cellular nanotherapeutic approach for retinal degenerative diseases. However, treatment specificity is compromised due to their high heterogeneity in size, content, functional effects, and parental cellular source. To improve this, engineered MSC-EXOs with increased drug-loading capacity, targeting ability, and resistance to bodily degradation and elimination have been developed. This review summarizes the recent advances in miRNAs of MSC-EXOs as a treatment for retinal degeneration, discussing the strategies and methods for engineering therapeutic MSC-EXOs. Notably, to address the single functional role of engineered MSC-EXOs, we propose a novel concept called "Compound Engineered MSC-EXOs (Co-E-MSC-EXOs)" along with its derived potential therapeutic approaches. The advantages and challenges of employing Co-E-MSC-EXOs for retinal degeneration in clinical applications, as well as the strategies and issues related to them, are also highlighted.

Periodontal Ligament Stem Cell-Derived Exosomes Regulate Muc5ac Expression in Rat Conjunctival Goblet Cells via Regulating Macrophages Toward an Anti-Inflammatory Phenotype.

BACKGROUND: Several studies have reported the protective effects of mesenchymal stem cell-derived exosomes (MSC-Exos) in reducing inflammation and decreasing conjunctival goblet cell (CGC) loss in dry eye disease. However, whether MSC-Exos provide anti-inflammatory profiles in macrophages, thus contributing to CGC protection, has remained elusive. METHODS: Macrophages were incubated with PKH26-labeled periodontal ligament mesenchymal stem cell-derived exosomes (PDLSC-Exos) for 12 h, and uptake of PDLSC-Exos by macrophages was observed by a confocal fluorescence microscope. The mRNA expression of TNF-α, IL-10, and Arg1 was detected by quantitative real-time polymerase chain reaction (qRT-PCR). The protein expression of TNF-α and IL-10 were quantified using western blotting. Then, CGCs were exposed to different macrophage supernatants and qRT-PCR was used to detect the Muc5ac mRNA expression of CGCs in response to or absence of cholinergic stimulation. ELISA was used to determine the Muc5ac secretion of CGCs in response to cholinergic stimulation. RESULTS: The uptake of PDLSC-Exos by M1 macrophages facilitates M2 macrophage polarization with the elevated expressions of IL-10 and Arg1. In macrophage supernatant-treated CGCs systems, PDLSC-Exo-treated M1 macrophage supernatant significantly enhanced the Muc5ac expression of CGCs in response to, or in the absence of, cholinergic stimulation, while the addition of PDLSC-Exos to the control macrophage supernatant did not generate a change in Muc5ac expression. Conversely, the addition of PDLSC-Exos to the diluted control macrophage supernatant induced a significant increase in Muc5ac expression. CONCLUSION: PDLSC-Exos could protect CGCs against M1 macrophage-mediated inflammation, and the protective effects of PDLSC-Exos are partly attributable to their effects on M1 macrophages.

The Use of Digital PCR for the Diagnosis of Demodex Blepharitis.

PURPOSE: This was a pilot study to evaluate the efficacy of digital polymerase chain reaction detection of Demodex in eyelid margin swabs for the diagnosis of Demodex blepharitis. This study aims to explore the possibility of digital polymerase chain reaction detection to improve the diagnostic accuracy of Demodex blepharitis detection. METHODS: Volunteers were prospectively recruited and classified by experienced doctors into suspected Demodex blepharitis or healthy controls using slit-lamp evaluation of the eyelid margin and an inquiry about symptoms. Three eyelashes from each eyelid were epilated from participants in each group for microscopic observation and mite counting. Then, swabs from the eyelid margins of each eye were collected after the eyelashes were epilated and stored at -80 °C for future DNA extraction and digital polymerase chain reaction detection. The positive or negative results of both methods were compared for diagnostic accuracy, and the Kappa value was also calculated to evaluate their consistency. RESULTS: The accuracy of the digital polymerase chain reaction detection was 71.6% and that of the mite counting method was 75%. Their combined accuracy was improved to 77.3%. The Kappa value of the two methods was 0.505, indicating moderate consistency. CONCLUSION: Digital polymerase chain reaction detection of Demodex from ocular surface swabs was painless and noninvasive and is a potentially accurate quantitative method available for diagnosing Demodex blepharitis. This method is also complementary to the conventional mite counting method, particularly when a sufficient number of eyelashes cannot be effectively epilated.

Dynamics of an epidemic model with general incidence rate dependent on a class of disease-related contact functions.

Starting from the idea of constructing the standard incidence rate, we take the effective contact times of individuals in the population per unit time as a contact function, $ T(\cdot) $, which depends on the population size. Considering the influence of disease on the contact function, the influence intensity factor of the disease affected by the infected person is integrated into the nonlinear incidence rate. We propose an epidemic model with a class of disease-related contact functions. Then, we analyze the well-posedness of the solutions of the model. By using the next generation matrix method, we get the basic reproduction number $ \mathcal{R}_0 $. We find that the existence and stability of the equilibria are not only related to $ \mathcal{R}_0 $, but also to the intensity of the disease affected for the infected person, $ \eta $, and the contact function, $ T(\cdot) $. We obtain some stability results under different assumptions about the contact function. Finally, we use MATLAB to simulate the system for several different contact functions. The numerical simulation results agree with our qualitative study. At the same time, we also prove that the system may have a Hopf bifurcation when the contact function $ T(\cdot) $ satisfies some corresponding conditions.

Tear film instability is associated with weakened colocalization between occludin and MUC5AC in scopolamine-induced dry eye disease (DED) rats.

PURPOSE: Dry eye disease (DED) is a disease with tear film instability because of multiple factors. This study was conducted to explore roles of occludin and MUC5AC in tear film instability in DED rat model. METHODS: A total of 20 SD rats were divided into DED group (n = 10) and normal control (NC) group (n = 10). DED rat model was established by subcutaneously injecting with scopolamine hydrobromide. Clinical examinations, including tear breakup time (tBUT), Schirmer's test and corneal fluorescein staining, were conducted to determine corneal functions. Transmission electron microscopy was used to measure the ultrastructures of corneal epithelial cells. Western blotting assay was used to identify occludin expression in corneal tissues of DED rats. Real-time PCR (RT-PCR) was performed to verify gene transcription of occludin and MUC5AC. Colocalization between occludin and MUC5AC was identified with confocal fluorescence microscopy. RESULTS: Tear breakup time was significantly shorter, and corneal fluorescein staining score was predominantly higher in DED rats compared to those in normal rats (P < 0.05). Normal rats showed a steady tear secretion throughout the whole experiments, while DED rats showed a dramatic reduction on day 14. DED rats demonstrated ultrastructural damage of Golgi apparatus and endoplasmic reticulum in corneal epithelial cells. Occludin and MUC5AC expressions were significantly downregulated in corneal tissue of DED rats compared with those of normal rats (P < 0.05). Percentage of occludin-MUC5AC-colocalized corneal epithelial cells in DED rats was significantly less compared with those in normal rats (P < 0.01). CONCLUSIONS: Tear film stability was damaged in scopolamine-induced DED rats because of the weakened colocalization between occludin and MUC5AC molecule. This study would provide a potential clue for the pathogenesis and a promising theoretical basis for clinical work of DED.

Vaccine failure, seasonality and demographic changes associate with mumps outbreaks in Jiangsu Province, China: Age-structured mathematical modelling study.

Measles, mumps and rubella (MMR) vaccine program was introduced in Jiangsu province of China in May 2008 and has been greatly contributed to decreasing of mumps cases. However, mumps has been resurging since May 2015. A number of studies have put forward that the resurgence of mumps is due to vaccine failure. In this paper, we investigated the other reasons for the resurging of mumps, such as the changes in seasonal transmission patterns and demographic structures, by using an age-structured mathematical model. We divided the history (January 2005 to May 2019) of mumps epidemics of Jiangsu province into three different stages: No vaccine stage (January 2005 to December 2008), effectively controlled stage (January 2009 to December 2014) and resurgence stage (January 2015 to May 2019). The features of mumps epidemics in three stages are compared under different demographic structures with same physical contact rate. The mumps transmission rate was increased in summer and dropped in November in stage III compared with that in stage I. The changes in demographic structures give a good explanation why the mumps outbreaked among children around 10 years old in stage I and around 5 years old in stage III. We have a conclusion that the vaccine failure, changes in seasonality and demographic structures were associated with the mumps outbreaks in recent years in Jiangsu province, China. We give the patterns of mumps dynamics considering age, vaccine, seasonality and demographic structures, which can help health program planners to implement more preventive interventions in mumps control during the period of higher risk of infection.

Comparison of 16S rDNA Amplicon Sequencing With the Culture Method for Diagnosing Causative Pathogens in Bacterial Corneal Infections.

PURPOSE: The purpose of this study was to explore if 16S rDNA amplicon sequencing can improve the conventional diagnosis of causative pathogens for bacterial corneal infection. METHODS: Corneal scraping and conjunctiva and eyelid margin swab samples from infected eyes of patients diagnosed with "bacterial corneal infection" and conjunctiva and eyelid margin swab samples from a random eye of healthy participants were collected. Each swab was used for both aerobic and anaerobic cultures and 16S rDNA amplicon sequencing. The V3 to V4 region of the 16S rDNA was amplified using polymerase chain reaction (PCR) and sequenced on the Illumina HiSeq 2500 Sequencing Platform. RESULTS: The overall culture positivity rate for all 72 samples was 69% (72% in the bacterial keratitis group and 67% in the healthy control group), whereas 1719 operational taxonomic units in total were generated using 16S rDNA amplicon sequencing with each sample showing 123 to 337 different genera. Staphylococcus, Corynebacterium, Propionibacterium, and Micrococcus most frequently appeared in culture, whereas Streptococcus, Acinetobacter, and Lactobacillus were the most common genera, with large ratios in 16S rDNA amplicon sequencing. The causative pathogens detected by the two methods were inconsistent for most samples, except for several corneal samples. CONCLUSIONS: We suggest that a combination of different techniques, such as clinical observation, microscopic analysis, culture, and next-generation sequencing techniques including 16S rDNA amplicon sequencing, should be used to comprehensively analyze pathogens in corneal and external ocular infections. TRANSLATIONAL RELEVANCE: This paper uses a basic research methodology for studying the microbiome in ocular samples to help improve the diagnostic accuracy of corneal and external ocular infections.

Corneal stromal mesenchymal stem cells: reconstructing a bioactive cornea and repairing the corneal limbus and stromal microenvironment.

Corneal stroma-derived mesenchymal stem cells (CS-MSCs) are mainly distributed in the anterior part of the corneal stroma near the corneal limbal stem cells (LSCs). CS-MSCs are stem cells with self-renewal and multidirectional differentiation potential. A large amount of data confirmed that CS-MSCs can be induced to differentiate into functional keratocytes in vitro, which is the motive force for maintaining corneal transparency and producing a normal corneal stroma. CS-MSCs are also an important component of the limbal microenvironment. Furthermore, they are of great significance in the reconstruction of ocular surface tissue and tissue engineering for active biocornea construction. In this paper, the localization and biological characteristics of CS-MSCs, the use of CS-MSCs to reconstruct a tissue-engineered active biocornea, and the repair of the limbal and matrix microenvironment by CS-MSCs are reviewed, and their application prospects are discussed.

Fiber reinforced GelMA hydrogel to induce the regeneration of corneal stroma.

Regeneration of corneal stroma has always been a challenge due to its sophisticated structure and keratocyte-fibroblast transformation. In this study, we fabricate grid poly (ε-caprolactone)-poly (ethylene glycol) microfibrous scaffold and infuse the scaffold with gelatin methacrylate (GelMA) hydrogel to obtain a 3 D fiber hydrogel construct; the fiber spacing is adjusted to fabricate optimal construct that simulates the stromal structure with properties most similar to the native cornea. The topological structure (3 D fiber hydrogel, 3 D GelMA hydrogel, and 2 D culture dish) and chemical factors (serum, ascorbic acid, insulin, and ß-FGF) are examined to study their effects on the differentiation of limbal stromal stem cells to keratocytes or fibroblasts and the phenotype maintenance, in vitro and in vivo tissue regeneration. The results demonstrate that fiber hydrogel and serum-free media synergize to provide an optimal environment for the maintenance of keratocyte phenotype and the regeneration of damaged corneal stroma.

Modeling and control of mosquito-borne diseases with Wolbachia and insecticides.

Mosquitoes cause more human suffering than any other organism. It is estimated that over one million people worldwide die from mosquito-borne diseases every year. With the continuous efforts of many researchers, Wolbachia gets more and more attention due to its characteristics of maternal transmission in mosquito population and it may cause cytoplasmic incompatibility (CI) which makes healthy females cannot fertilize normally after mating with infected males. In this paper, mathematical models are established to study Wolbachia transmission in mosquito population, and integrated mosquito control strategies are explored. Firstly, a classical ordinary differential system with general birth and death rate functions is established to describe the maternal transmission and CI effect. It is shown that the replacement strategy that the Wolbachia-uninfected mosquitoes are replaced by the infected ones is determined by the initial infection frequency. And Wolbachia spreads more easily for greater maternal transmission and CI rate. Moreover, all the wild mosquitoes will eventually be infected with Wolbachia if the maternal transmission is complete. Secondly, an impulsive state feedback control model is constructed to describe the integrated mosquito control. Besides Wolbachia, insecticides are sprayed when the quantity of mosquitoes reaches some Economic Threshold. The existence and stability of Wolbachia replacement periodic solution are discussed. Finally, some discussions are done and the future research directions are prospected.

A Host-Parasite System with Multiple Parasite Strains and Superinfection Revisited: The Global Dynamics.

In this paper, we revisit a host-parasite system with multiple parasite strains and superinfection proposed by Nowak and May (Proc R Soc Lond B 255(1342):81-89, 1994), and study its global dynamics when we relax the two strict conditions assumed therein. As for system with two parasite strains, we derive that the basic reproduction number [Formula: see text] is the threshold condition for parasite extinction and the invasion reproduction number [Formula: see text] is the subthreshold condition for coexistence of two parasite strains. As for system with three parasite strains, we are surprised to discover the global stability of parasite-free and coexistence equilibrium, which is distinct from the previous result. Furthermore, for system with n strains, we obtain the global asymptotical stability of the parasite-free equilibrium, conjecture a general result on the global stability of coexistence equilibrium and provide two numerical examples to testify our conjecture.

Expression of visual system homeobox 1 in human keratoconus.

AIM: To investigate the expression of visual system homeobox 1 (VSX1) and myofibroblast marker alpha smooth muscle actin (α-SMA) in keratoconus (KC). METHODS: Thirty corneal tissue were collected from KC patients after corneal transplantation and 15 normal donor corneas were obtained. All corneal tissues divided into 4 parts for different detections. Scanning electron microscopy was used to observe the ultrastructure of the specimens. VSX1 and α-SMA localization in cornea tissues was detected using immunofluorescence histochemistry. Reverse transcription-quantitative polymerase chain reaction (RT-qPCR) and Western blot were performed to analyze the expression level of VSX1 and α-SMA. RESULTS: Compared to normal cornea tissue, the collagen fibers in KC stroma were distortional and attenuated and keratocytes were abnormally changed. VSX1 and α-SMA located in the corneal stroma. The mRNA and protein expression level of VSX1 in KC were about 3 times as high as that of normal tissue (P<0.001). α-SMA was hardly expressed in the normal corneas, however, its expression in the KC was about 1.5 times higher than that of the normal corneas (P<0.0001). CONCLUSION: Compared with normal corneal the expression of VSX1 and α-SMA in KC both increased. VSX1 is related to the activation of keratocytes and involved in the pathogenesis of keratoconus.

A sex-structured model with birth pulse and release strategy for the spread of Wolbachia in mosquito population.

Dengue fever is one of the most important diseases causing illness and death all over the world, which brings tremendous threat to peoples' life and property security, especially in the undeveloped areas. The main vector, Aedes aegypti, must be controlled to prevent the transmission of dengue. There are a variety of methods to control it. Wolbachia is an innovative bacterium which breaks the dengue transmission cycle for its characteristics of cytoplasmic incompatibility and maternal transmission. In this paper, a sex-structured model with birth pulse is established to study the spread of Wolbachia in mosquito population. The results show that if the maternal transmission is perfect, Wolbachia will spread successfully. Moreover, all the mosquitoes will be infected with Wolbachia. If the maternal transmission is imperfect, there are two locally asymptotically stable periodic solutions. One is Wolbachia-extinction periodic solution, and the other is part replacement periodic solution. Numerical simulations show that the initial occupancy of Wolbachia-infected mosquitoes has an important effect on the success of part replacement strategy. If the initial occupancy is relatively large, the part replacement strategy can be successful. Furthermore, in consideration of the fact that the initial occupancy cannot be always large enough in the wild nature, to release Wolbachia-infected mosquitoes artificially into the wild nature becomes necessary. Therefore, we add a release strategy into the sex-structured model with birth pulse for further analysis. The condition to ensure the stability of the Wolbachia total replacement periodic solution is obtained. Finally, the effect of the release quantity is simulated numerically.

A reaction-diffusion within-host HIV model with cell-to-cell transmission.

In this paper, a reaction-diffusion within-host HIV model is proposed. It incorporates cell mobility, spatial heterogeneity and cell-to-cell transmission, which depends on the diffusion ability of the infected cells. In the case of a bounded domain, the basic reproduction number [Formula: see text] is established and shown as a threshold: the virus-free steady state is globally asymptotically stable if [Formula: see text] and the virus is uniformly persistent if [Formula: see text]. The explicit formula for [Formula: see text] and the global asymptotic stability of the constant positive steady state are obtained for the case of homogeneous space. In the case of an unbounded domain and [Formula: see text], the existence of the traveling wave solutions is proved and the minimum wave speed [Formula: see text] is obtained, providing the mobility of infected cells does not exceed that of the virus. These results are obtained by using Schauder fixed point theorem, limiting argument, LaSalle's invariance principle and one-side Laplace transform. It is found that the asymptotic spreading speed may be larger than the minimum wave speed via numerical simulations. However, our simulations show that it is possible either to underestimate or overestimate the spread risk [Formula: see text] if the spatial averaged system is used rather than one that is spatially explicit. The spread risk may also be overestimated if we ignore the mobility of the cells. It turns out that the minimum wave speed could be either underestimated or overestimated as long as the mobility of infected cells is ignored.

Modelling the Transmission Dynamics and Control of Mumps in Mainland China.

Mumps is a common childhood viral disease and children have been vaccinated throughout the world since 1967. The incidence of mumps has increased with more than 300,000 young people infected with mumps annually in mainland China since 2005. Therefore, we designed and analyzed long-term mumps surveillance data in an SVEILR (susceptible-vaccinated-exposed-severely infectious-mildly infectious-recovered) dynamic transmission model with optimized parameter values to describe the dynamics of mumps infections in China. There were 18.02% of mumps infected young adults seeking medical advice. The vaccine coverage has been insufficient in China. Young adults with frequent contact and mild infection were identified as a major driver of mumps epidemics. The reproduction number of mumps was determined 4.28 in China. Sensitivity analysis of the basic reproduction number and the endemic equilibrium was conducted to evaluate the effectiveness of mumps control measures. We propose to increase the vaccine coverage and make two doses of MMR (Measles, mumps and rubella) vaccines freely available in China.

Corneal collagen cross-linking and liposomal amphotericin B combination therapy for fungal keratitis in rabbits.

AIM: To observe the therapeutic effect of corneal collagen cross-linking (CXL) in combination with liposomal amphotericin B in fungal corneal ulcers. METHODS: New Zealand rabbits were induced fungal corneal ulcers by scratching and randomly divided into 3 groups, i.e. control, treated with CXL, and combined therapy of CXL with 0.25% liposomal amphotericin B (n=5 each). The corneal lesions were documented with slit-lamp and confocal microscopy on 3, 7, 14, 21 and 28d after treatment. The corneas were examined with transmission electron microscopy (TEM) at 4wk. RESULTS: A rabbit corneal ulcer model of Fusarium was successfully established. The corneal epithelium defect areas in the two treatment groups were smaller than that in the control group on 3, 7, 14 and 21d (P<0.05). The corneal epithelium defect areas of the combined group was smaller than that of the CXL group (P<0.05) on 7 and 14d, but there were no statistical differences on 3, 21 and 28d. The corneal epithelium defects of the two treatment groups have been healed by day 21. The corneal epithelium defects of the control group were healed on 28d. The diameters of the corneal collagen fiber bundles (42.960±7.383 nm in the CXL group and 37.040±4.160 nm in the combined group) were thicker than that of the control group (24.900±1.868 nm), but there was no difference between the two treatment groups. Some corneal collagen fiber bundles were distorted and with irregular arrangement, a large number of fibroblasts could be seen among them but no inflammatory cells in both treatment groups. CONCLUSION: CXL combined with liposomal amphotericin B have beneficial effects on fungal corneal ulcers. The combined therapy could alleviate corneal inflammattions, accelerate corneal repair, and shorten the course of disease.

Acellular ostrich corneal stroma used as scaffold for construction of tissue-engineered cornea.

AIM: To assess acellular ostrich corneal matrix used as a scaffold to reconstruct a damaged cornea. METHODS: A hypertonic saline solution combined with a digestion method was used to decellularize the ostrich cornea. The microstructure of the acellular corneal matrix was observed by transmission electron microscopy (TEM) and hematoxylin and eosin (H&E) staining. The mechanical properties were detected by a rheometer and a tension machine. The acellular corneal matrix was also transplanted into a rabbit cornea and cytokeratin 3 was used to check the immune phenotype. RESULTS: The microstructure and mechanical properties of the ostrich cornea were well preserved after the decellularization process. In vitro, the methyl thiazolyl tetrazolium results revealed that extracts of the acellular ostrich corneas (AOCs) had no inhibitory effects on the proliferation of the corneal epithelial or endothelial cells or on the keratocytes. The rabbit lamellar keratoplasty showed that the transplanted AOCs were transparent and completely incorporated into the host cornea while corneal turbidity and graft dissolution occurred in the acellular porcine cornea (APC) transplantation. The phenotype of the reconstructed cornea was similar to a normal rabbit cornea with a high expression of cytokeratin 3 in the superficial epithelial cell layer. CONCLUSION: We first used AOCs as scaffolds to reconstruct damaged corneas. Compared with porcine corneas, the anatomical structures of ostrich corneas are closer to those of human corneas. In accordance with the principle that structure determines function, a xenograft lamellar keratoplasty also confirmed that the AOC transplantation generated a superior outcome compared to that of the APC graft.

Establishment of a novel therapeutic vector targeting the trigeminal ganglion in rats.

BACKGROUND: In the pathogenesis of herpes simplex keratitis, herpes simplex virus type 1 (HSV-1) infection begins in corneal epithelium cells and then progresses through the sensory nerve endings and finally travels up forward to the trigeminal ganglion (TG), where it remains as latent virus. The available anti-HSV therapies do not completely suppress the recurrence of active HSV-1 infection. The aim of this study was to establish a novel replication-defective (rd) HSV-1 (rdHSV) vector (rdHSV-interferon gamma [IFNγ]) that could effectively target the TG. METHODS: Recombinant HSV-1 virus was inserted into a shuttle plasmid carrying IFNγ to establish the rdHSV-IFNγ vector. Safety was evaluated in vitro by 50% cellular cytotoxicity in transfected SH-SY5Y neuroblastoma cells and in vivo by Kaplan-Meier survival estimate and infection rate. Wistar rats were immunized with rdHSV-IFNγ to evaluate the TG targeting efficiency. Real-time polymerase chain reaction and Western blot assays were used to evaluate IFNγ mRNA and protein expression and rdHSV-IFNγ localization. RESULTS: The rdHSV-IFNγ vector was successfully constructed and showed high in vitro safety and overall survival and a corneal infection rate similar to that of control rats immunized with saline (control group; P>0.05). Real-time polymerase chain reaction and immunohistochemistry assays confirmed IFNγ expression and effective TG targeting on days 14 and 21, which increased with postimmunization time. Moreover, IFNγ was expressed sufficiently in the TG tissues. CONCLUSION: The rdHSV-IFNγ can act as an effective gene transporting vector that carries the therapeutic genes to the TG and triggers its expression.