The Species Composition of Microbiota and Biological Properties of Dominant Bacterial Species in the Lacrimal Fluid in Patients with Endogenous Uveites

The purpose: to study the species composition of the lacrimal fluid microbiota in patients with endogenous uveitis and the biological properties of dominant species of bacteria.Patients and Methods. A bacteriological study of the lacrimal fluid of 107 patients (114 eyes) with endogenous uveitis and 28 practically healthy persons (control) was carried out. The research was performed using the conventional methods. The isolated microorganisms were identified up to the species. We studied the main biological properties of bacteria — the presence of persistence and virulence factors of isolated dominant species.The results. 43 strains of various types of bacteria were isolated from the lacrimal fluid of patients with uveitis, bacteria of the genus Staphylococcus prevailed, S. aureus was isolated in 23.3 % of cases, and the remaining strains were representatives of coagulase-negative staphylococci. The species composition of the bacteria isolated from practically healthy individuals did not differ in essence. Isolated strains of S. aureus had a significant set of virulence factors. All isolates lysed sheep red blood cells, 80 % of the cultures had lecithovitellase and 60 % — DNA-se activity. Representatives of coagulase-negative staphylococci were characterized only by hemolytic activity. In the study of persistence factors, it was found that all the studied strains of S. aureus and the vast majority of representatives of coagulase-negative staphylococci isolated from tears were resistant to lysozyme. Biofilm formation was more typical for S. aureus strains 90 % versus 34.4 % for representatives of coagulase-negative staphylococci (t = 2.194, p = 0.034). Similar patterns were found in relation to complement resistance, 90 % of S. aureus cultures and 37.5 % of coagulase-negative staphylococci (t = 2.055; p = 0.046).Conclusion. Among the microorganisms isolated from the lacrimal fluid, Staphylococcus bacteria prevailed, from which S. cohnii was most often isolated. S. aureus to a greater extent than coagulase-negative staphylococci had a certain set of virulence and persistence factors. The microbiocenosis of the lacrimal fluid of patients with endogenous uveitis did not differ from practically healthy individuals. The presence of S. aureus in lacrimal fluid is a risk factor for the development of endogenous uveitis in cases of chronic oral and ENT organ diseases.

Immunological aspects of bacterial keratites in patients with diabetes mellitus

Background. The purpose was to analyze the cytokine balance of lacrimal fluid in patients with bacterial keratitis and diabetes mellitus (DM) at the first visit and to identify the immunological aspects of the disease. Materials and methods. The analysis of pro- and anti-inflammatory cytokine concentration in the lacrimal fluid was performed in 17 patients with type 1 DM and bacterial keratitis and 15 nondiabetic patients with bacterial keratitis at the first visit. Data from 14 healthy individuals were used for comparison. In addition to standard ones, ophthalmic examination methods included bacteriological examination, fluorescein test, anterior segment optical coherence tomography, non-contact corneal aesthesiometry. The levels of interleukin (IL) 1β, IL-6 and IL-10 in the lacrimal fluid of the sick and the contralateral eye were determined by a quantitative colorimetric enzyme-linked immunosorbent assay using ELISA kits. Results. In DM patients with bacterial keratitis, the concentration of IL-1β and IL-6 in the lacrimal fluid of the sick eye exceeded that in healthy individuals (p < 0.05) and did not differ significantly from nondiabetic patients with bacterial keratitis (p > 0.05). In the lacrimal fluid of the contralateral eye of DM patients with bacterial keratitis, the level of IL-1β and IL-6 exceeded the corresponding indicators of nondiabetic patients with bacterial keratitis and healthy individuals (p < 0.05). The concentration of IL-10 in the lacrimal fluid of the contralateral eye in DM patients with bacterial keratitis exceeded that in healthy individuals (p < 0.05) and did not significantly differ from those in nondiabetic patients with bacterial keratitis (p > 0.05). Conclusions. DM patients with bacterial keratitis have immunological features of the disease.

PECULIARITIES OF THE LACRIMAL FLUID CYTOKIN PROFILE IN PATIENTS WITH BACTERIAL KERATITITIS OF THE II DEGREE SEVERETY AND DIABETES MELLITUS

Summary. The aim was to identify the features of the lacrimal fluid cytokine profile in patients with bacterial keratitis of the II degree of severity and type 1 DM in the dynamics of observation. Materials and methods. The lacrimal fluid cytokine profile was analyzed in 6 patients with type 1 DM and bacterial keratitis of the II degree of severity and 6 nondiabetic patients with bacterial keratitis of the II degree of severity on the 1st day, 10-14th day and 24th day of observation. Data from 14 healthy individuals of the appropriate age were used for comparison. II degree of severity of bacterial keratitis was determined according to our proposed scheme at a sum of points 15-21. All patients were treated with instillations of ofloxacin, antiseptics, antioxidants, reparents, artificial tears, mydriatics, systemic anti-inflammatory therapy. The concentration of interleukins (IL)-1β, IL-6 and IL-10 in the lacrimal fluid of the sick and the contralateral eye was determined by quantitative colorimetric enzyme-linked immunosorbent assay using ELISA kits. Results and discussion. In patients with bacterial keratitis of the II degree of severity, both with DM and nondiabetic, within 10-14 days of observation invented the expression of proinflammatory cytokines IL-1β and IL-6 in the lacrimal fluid of the sick eye and anti-inflammatory cytokine IL-10 - in lacrimal fluid of the contralateral eye (p<0.05). In DM patients, a longer expression of IL-1β and IL-6 was detected in the lacrimal fluid of the sick and the contralateral eye, which was observed on the 24th day of the study (p<0.05). Conclusions. There are features of the lacrimal fluid cytokine profile in DM patients with bacterial keratitis of the II severity in the dynamics of observation, namely the longer expression of proinflammatory cytokines IL-1β, IL-6 in the sick and the contralateral eye.

Balance of Drug Residence and Diffusion in Lacrimal Fluid Determine Ocular Bioavailability in In Situ Gels Incorporating Tranilast Nanoparticles

We previously designed ophthalmic formulations (nTRA) containing tranilast nanoparticles (Tra-NPs) with high uptake into ocular tissues. In this study, we used in situ gel (ISG) bases comprising combinations of pluronic F127 (F127) and methylcellulose (MC/F127), pluronic F68 (F68/F127), and Carbopol (Car/F127), and we developed in situ gels incorporating Tra-NPs (Tra-NP-incorporated ISNGs) such as nTRA-F127, nTRA-MC/F127, nTRA-F68/F127, and nTRA-Car/F127. Moreover, we demonstrated the therapeutic effect on conjunctival inflammation using lipopolysaccharide-induced rats. Each Tra-NP-incorporated ISNG was prepared by the bead mill method, the particle size was 40–190 nm, and the tranilast release and diffusion from formulation were nTRA > nTRA-F127 > nTRA-F68/F127 > nTRA-Car/F127 > nTRA-MC/F127. In the Tra-NP-incorporated ISNGs, the tranilast residence time in the lacrimal fluid, cornea, and conjunctiva was prolonged, although the Cmax was attenuated in comparison with nTRA. On the other hand, no significant difference in conjunctival inflammation between non- and nTRA-F127-instilled rats was found; however, the nTRA-F68/F127, nTRA-Car/F127, and nTRA-MC/F127 (combination-ISG) attenuated the vessel leakage, nitric oxide, and tumor necrosis factor-α expression. In particular, nTRA-F68/F127 was significant in preventing the conjunctival inflammation. In conclusion, we found that the combination-ISG base prolonged the residence time of Tra-NPs; however, Tra-NP release from the formulation was attenuated, and the Tmax was delayed longer than that in nTRA. The balance of drug residence and diffusion in lacrimal fluid may be important in providing high ocular bioavailability in formulations containing solid nanoparticles.

IMMUNOLOGICAL FEATURES OF THE DYNAMICS OF THE I DEGREE SEVERETY BACTERIAL KERATITES DEPENDING ON THE PRESENCE OF DIABETES MELLITUS

Summary. In patients with diabetes mellitus (DM) there are features of the dynamics of the I degree severity bacterial keratitis, leading to prolongation of the disease and worsening of its consequences. The aim was to identify immunological features of the dynamics of the I degree severity bacterial keratitis depending on the presence of type 1 DM. Materials and methods. The level of interleukins (IL) was analyzed, namely IL-1β, IL-6 and IL-10 in the lacrimal fluid of the sick and the contralateral eye of 6 patients with type 1 DM and the I degree severity bacterial keratitis and 6 nondiabetic patients with I degree severity bacterial keratitis on the 1st day, 10-14th day and 24th day of observation. Data from 14 healthy individuals of the appropriate age were used for comparison. The severity of bacterial keratitis was determined according to our scheme at a total score of ≤ 14. All patients were treated with instillations of ofloxacin, antiseptics, antioxidants, reparents, artificial tears, mydriatics, systemic anti-inflammatory therapy. The level of IL in the lacrimal fluid was determined by quantitative colorimetric enzyme-linked immunosorbent assay using ELISA kits by ANOGEN according to the instructions for use. Results and discussion. In patients with I degree severity bacterial keratitis, both DM and nondiabetic, on the 1st day of observation there was an increase in the level of IL-1β and IL-6 in the lacrimal fluid of the sick eye and IL-10 – of the contralateral eye compared to healthy persons, on the 10th-14th day — increase in the level of IL-1β and IL-6 of the sick and the contralateral eye, as well as IL-10 – of the contralateral eye (p<0.05). On the 24th day in nondiabetic patients with I degree severity bacterial keratitis normalization of interleukin levels in the lacrimal fluid of the sick and the contralateral eye was found, but in DM patients remained elevated levels of IL-1β, IL-6, both in the sick and in the contralateral eye (p<0.05). Conclusions. In DM patients there are immunological features of the dynamics of the I degree severity bacterial keratitis, namely a longer increase in the level of proinflammatory cytokines IL-1β, IL-6 in the lacrimal fluid of the sick and the contralateral eye.

Melatonin level as a risk factor for age-related macular degeneration

Background. The current trend towards an increase in age-related macular degeneration (AMD) incidence rate in the population, including the working-age population, with a possible loss of professional activity, indicates the need for early preclinical identification of risk groups, timely prevention and treatment. In the prevention and treatment of AMD, the prospect of using melatonin is being actively discussed.Aim: to analyze serum and tear fluid levels of melatonin in patients with AMD and study their correlation with risk factors.Materials and methods. In the course of the study, two groups were formed: the main group - patients with non-exudative AMD and senile cataract (n = 40) and the reference group - conditionally healthy patients without AMD and cataract (n = 20). Patients of both groups were surveyed to identify risk factors for AMD. The content of melatonin in blood serum and lacrimal fluid was determined by enzyme-linked immunosorbent assay using the Melatonin ELISA Kit (USA).Results. In the course of the study, it was found that the concentration of melatonin in blood serum and tear fluid in patients with AMD was significantly lower than in patients of the reference group (p <0.05). Serum and tear fluid melatonin levels depend on the following factors: age, body mass index (BMI), arterial hypertension, eye color, insomnia, and night work. It is possible that the local determination of melatonin in the lacrimal fluid can be a biomarker in the determination of ophthalmic pathological conditions.Conclusion. The obtained results can be used as recommendations for clarifying individual regimens for the use of melatonin, especially in the treatment of patients with AMD.

Submicroscopic characteristics of the lacrimal fluid in vitreomacular traction syndrome, diabetic retinopathy compared with the norm

Study of the lacrimal fluid (LF) as a constant microenvironment of the anterior part of the eye which is the only atraumatically accessible substrate for the diagnosis and research of ophthalmic diseases, such as vitreomacular traction syndrome (VMTS), diabetic retinopathy (DR), makes it possible to study it using electronic microscopy methods. All studied LF samples contain cells and cell fragments; exosomes which are vesicles (40–100 nm) localized in multivesicular bodies, transmitting signals between cells and carrying markers of many diseases. Analysis of the samples revealed changes in the occurrence of these structures in VMTS and DR in comparison with healthy subjects. In this work, the components of the LF were visualized and their changes were established in DR and VMTS, which proves the value of the LF as a diagnostic substrate and determines the need for further research in order to formulate clear criteria for the diagnosis of these diseases in the early stages. Key words: lacrimal fluid; electronic microscopy; vitreomacular traction syndrome; diabetic retinopathy.

Quantitative Assessment of Age and Gender Related Changes in Human Lacrimal Fluid Composition in Subjects

Purpose: The frequency of eyelids blink rate has effect on the integrity of the tear fluid and replacement of old of tears on the ocular surface. Comprehension of the effect of advancing age and gender on tears fluid stability and quality is contentious. Basal tears continuously bathe the ocular facet lubricating and providing protection from viral and bacterial infections, cooling the surface as warmth encourages pathogen. Basal tear dysfunction basically results in conjunctival and corneal conditions which exposes the ocular facet to dryness, ulceration, opacity and even blindness. This study determined the quantitative assessment of age and gender related changes in human lacrimal fluid composition in subjects Methods: The design involved 400 (192 males and 208 females) human subjects aged 6 years and above who do not wear contact lenses, not on topical medication and having healthy eyes grouped into twelve groups based on age and gender. Maximum blink interval (for tear break-up time) and phenol red thread procedure (for tear volume) were carried out monocularly for each subject. Results: Findings from this study have established that tear fluid stability among different age groups; are often higher for group 6-12years and lower for 60years and above; therefore, study established that tear fluid stability is age dependent. In addition, tear fluid stability (average MBI. Tear volume and flow rate) are gender independent. Conclusion: The present study revealed that tears fluid breakup time increases with decrease in tears fluid flow rate, volume and flow.

The Role of Biologically Active Aqueous Humor Molecules of the Anterior Chamber and Lacrimal Fluid in the Implementation of the Hypotensive Effect of Non-Penetrating Deep Sclerectomy

To date, the factors affecting the course of the reparative process after non-penetrating deep sclerectomy (NPDS) have not been fully determined. There is no systematic information about the regulatory role of the cytokines TGF-β, IL-6, IL-8 and MMP-9, VEGF A 121 and 165 in the formation mechanisms of the newly created pathways consistency of intraocular aqueous humor outflow.Purpose. To determine possible ways of impact of biologically active aqueous humor molecules of the anterior chamber and lacrimal fluid on the hypotensive effect of non-penetrating deep sclerectomy.Methods. A prospective study of 65 patients with open-angle glaucoma before and 12 months after NPDS and 22 patients without eye hydrodynamic disorders with the determination of the initial concentrations of biologically active molecules in the lacrimal fluid and aqueous humor of the anterior chamber. Twelve months after NPDS all patients were divided into three groups, depending on the hypotensive effect of the operation, according to the criteria.Results. Multivariate discriminant analysis showed the greatest inter-group differences, calculated by the square of the Mahalanobis distance, between group 3 with no hypotensive effect of NPDS and the control group (R2 = 8.48, p = 0.001). The most informative features that determine the differences between the 4 groups in the total population, calculated according to the Fischer F-test, were MMP-9 (F = 14.7, p = 0.001) and TGF-β (F = 7.08, p = 0.001) in the aqueous humor of the anterior chamber. In pairwise comparison of groups 1 and 2, the maximum level of significance according to the F-criterion was characteristic of the level of tear IL-6 (F = 21.25, p = 0.001), with approximately equal degree – IL-8 (F = 7.85, p = 0.001) and VEGF (F = 7.12, p = 0.001), to a lesser extent TGF of aqueous humor (F = 4.43, p = 0.001) and MMR-9 (F = 2.23, p = 0.001). Between groups 1–3, the maximum differences according to the Fisher criterion were observed in the IL-8 (F = 20.99, p = 0.001), TGF (F = 8.75, p = 0.001) and to a lesser extent – TGF (F = 5.83, p = 0.001).Conclusion. The analysis of the obtained data showed the decisive role of the imbalance of proinflammatory cytokines, signaling proteins with prolymphoangiogenic activity, and MMP-9 in the aqueous humor of the anterior chamber, as well as in the initial state of the lacrimal fluid in the postoperative healing processes after NPDS.

Continuous Non-Invasive Glucose Monitoring via Contact Lenses: Current Approaches and Future Perspectives

Diabetes mellitus (DM) is a chronic disease that must be carefully managed to prevent serious complications such as cardiovascular disease, retinopathy, nephropathy and neuropathy. Self-monitoring of blood glucose is a crucial tool for managing diabetes and, at present, all relevant procedures are invasive while they only provide periodic measurements. The pain and measurement intermittency associated with invasive techniques resulted in the exploration of painless, continuous, and non-invasive techniques of glucose measurement that would facilitate intensive management. The focus of this review paper is the existing solutions for continuous non-invasive glucose monitoring via contact lenses (CLs) and to carry out a detailed, qualitative, and comparative analysis to inform prospective researchers on viable pathways. Direct glucose monitoring via CLs is contingent on the detection of biomarkers present in the lacrimal fluid. In this review, emphasis is given on two types of sensors: a graphene-AgNW hybrid sensor and an amperometric sensor. Both sensors can detect the presence of glucose in the lacrimal fluid by using the enzyme, glucose oxidase. Additionally, this review covers fabrication procedures for CL biosensors. Ever since Google published the first glucose monitoring embedded system on a CL, CL biosensors have been considered state-of-the-art in the medical device research and development industry. The CL not only has to have a sensory system, it must also have an embedded integrated circuit (IC) for readout and wireless communication. Moreover, to retain mobility and ease of use of the CLs used for continuous glucose monitoring, the power supply to the solid-state IC on such CLs must be wireless. Currently, there are four methods of powering CLs: utilizing solar energy, via a biofuel cell, or by inductive or radiofrequency (RF) power. Although, there are many limitations associated with each method, the limitations common to all, are safety restrictions and CL size limitations. Bearing this in mind, RF power has received most of the attention in reported literature, whereas solar power has received the least attention in the literature. CLs seem a very promising target for cutting edge biotechnological applications of diagnostic, prognostic and therapeutic relevance.