Evaluation of corneal epithelial thickness mapping using anterior segment OCT in children with vernal keratoconjunctivitis.

PURPOSE: To evaluate the corneal epithelial thickness by anterior segment OCT in children with vernal keratoconjunctivitis (VKC) compared to normal participants. PATIENT AND METHODS: A cross-sectional case control observational study was conducted on children aged from 5 to 18 years with VKC. The study included 142 eyes divided into two groups: 71 eyes of VKC group and 71 eyes of age matched control group. Anterior segment OCT epithelial mapping for the central 5-mm was done to evaluate epithelial thickness-based variables. RESULTS: Corneal epithelial thickness mapping showed significant superior thinning (51.07 ± 4.11) µm in VKC group compared to controls (52.54 ± 2.01) µm (p = 0.008), a decreased MIN epithelial thickness (45.99 ± 6.52) µm compared to controls (50.11 ± 1.91) µm (p < 0.001) and more negative (MIN-MAX) value (- 11.77 ± 9.38) indicating focal epithelial thinning compared to (- 5.80 ± 1.88) in controls (p = 0.001). In palpebral VKC, MIN epithelial thickness showed significant thinning (48. 38 ± 3.45) µm compared to controls (50.11 ± 1.91) µm (p = 0.001). Eyes with mixed VKC showed superior epithelial thinning (50.70 ± 4.59) µm compared to (52.54 ± 2.01) µm in controls (p = 0.025) and significant decreased MIN epithelial thickness (43.78 ± 7.83) µm compared to (50.11 ± 1.91) µm in control group. CONCLUSIONS: VKC is a disease primarily affecting the corneal epithelium. Corneal epithelial thickness mapping may be considered to assess the integrity of the ocular surface in eyes with VKC, and to detect corneal epithelial changes. Disease phenotype may influence the corneal epithelial changes, and the disease duration is another factor influencing these changes.

Rapid back flushed direct sample injection bio-analytical HPLC-UV method for therapeutic drug monitoring of terbinafine.

A rapid back flushed (BF) direct sample injection (DSI) high-performance liquid chromatography (HPLC) with UV detection (BF-DSI-HPLC-UV) has been developed to determine terbinafine (TERB) in human serum. For online solid phase extraction step, an isocratic mobile phase of phosphate buffer saline (pH 7.4) at 1 mL/min and a short protein-coated ODS column (PC-ODS-column) were used for the purification and enrichment of TERB. Two different chromatographic modes of PC-ODS-column were simultaneously operated. Macromolecular proteins were extracted by size-exclusion liquid chromatography, while TERB trapping and enrichment were achieved through reversed-phase liquid chromatography. The clear fraction containing TERB was transferred from the PC-ODS-column by BF mode onto the quantification step through a high pressure switching valve. An analytical mobile phase consisting of 80% methanol and 1% triethylamine in distilled deionized water (pH) 6 at 1 mL/min was used for the final separation on an ODS analytical column. TERB was quantified and detected by UV-detector at 224 nm. The proposed method showed high correlation coefficient (>0.999) over the concentrations range 4-1600 ng/mL with recoveries ranging from 98.48 to 93.86%. Measurement of TERB concentration in serum after administration of a single dose of 250 mg oral tablet was used to evaluate the applicability of the BF-DSI-HPLC-UV for pharmacokinetic study.

Bio-analytical liquid chromatographic-based method with a mixed mode online solid phase extraction for drug monitoring of fluconazole in human serum.

A simple, cost-effective and sensitive liquid chromatography-based bio-analytical method has been developed and validated for therapeutic drug monitoring of fluconazole (FLUC) in human serum. Integration of online mixed-mode solid-phase extraction (SPE) into the analytical system was the key for direct injection of untreated serum samples. A short protein-coated (PC) µBondapak CN silica column (PC-µB-CN-column) as a SPE tool and phosphate buffer saline (PBS) (pH 7.4) as an eluent were applied in the extraction step. PC-µB-CN-column operates in two different chromatographic modes. Using PBS, proteins were extracted from serum samples by size-exclusion liquid chromatography, while FLUC trapping was reversed-phase liquid chromatography dependent. FLUC was then eluted from the PC-µB-CN-column onto the quantification position using a mixture of acetonitrile-distilled deionized water (20:80, v/v) as an eluent and ODS analytical column. FLUC was separated at ambient temperature (22 ± 1 °C) and detected at 260 nm. The method was linear over the range of 200-10000 ng/mL. FLUC recovery in untreated serum samples ranged from 97.8 to 98.8% and showed good accuracy and precision. The reliability of the developed method was evaluated by studying the pharmacokinetic profile of FLUC in humans after an oral administration of a single 150 mg tablet.

Direct infusion nano-electrospray ionization mass spectrometry for therapeutic drug monitoring of ciprofloxacin and its metabolites in human saliva.

A rapid and sensitive method based on direct infusion-nano-electrospray ionization mass spectrometry (DI-nESI-MS) has been developed for the detection and quantification of ciprofloxacin and its metabolites in human saliva. Saliva samples were collected after the oral administration of 500 mg ciprofloxacin tablets. Internal standard (IS), tamoxifen, was added to the collected samples, and then diluted with the ionization solvent, centrifuged and filtered. An aliquot of 4 µL of the filtrate was loaded into a nanospray (NS) capillary. The NS capillary was then fitted into an off-line ion source and the instrument was operated to acquire a two-minute run by applying a voltage of 1000 V (positive-ion detection mode). Quantification of ciprofloxacin relied on the ratio of its peak intensity to the IS peak intensity. The DI-nESI-MS method was validated and provided satisfactory precision with relative standard deviation ranging from 0.39 to 7.48 % and accuracy with relative error ranging from -2.12 to 9.72 %. The calibration curve showed good linearity (r2) > 0.999 over the concentration range of 10-4000 ng/mL. These results verify the effectiveness of the DI-nESI-MS method for monitoring of ciprofloxacin and its metabolites in human saliva samples.

Dilute-and-shoot-based direct nano-electrospray ionization tandem mass spectrometry as screening methodology for multivitamins in dietary supplement and human urine.

INTRODUCTION: In recent years, analytical screening methods for simultaneous detection of multivitamins have gained substantial attention to ensure quality and public confidence in dietary supplements. Even so, few analytical methods have been proposed for simultaneous analysis of multivitamin constituents due to the large divergence in chemical characteristics. OBJECTIVES: In the present study, the objective was to develop a simple and rapid direct nano-electrospray ionization-tandem mass spectrometry (DI-nano-ESI-MS/MS) method for targeted detection of water soluble vitamins, fat soluble vitamins, amino acids, royal jelly, ginkgo biloba, and ginseng in a dietary supplement. The applicability of dilute-and-shoot-based DI-nano-ESI-MS/MS to analyze the same tested compounds and their related metabolites in clinical samples was also examined. METHODS: Intact urine mixed with the ionization solvent was loaded (4-µL aliquot) into a nanospray (NS) capillary of 1-µm tip diameter. The NS capillary was then fitted into an off-line ion source at a distance of 5 mm from MS aperture. The sample was directly injected by applying a voltage of 1.1 kV, producing a numerous of m/z peaks for analysis in mere minutes. RESULTS: The DI-nano-ESI-MS/MS method successfully identified almost all dietary supplement components, as well as a plethora of component-related metabolites in clinical samples. In addition, a new merit of the proposed method for the detection of index marker and chemical contaminants as well as subspecies identification was investigated for further quality evaluation of the dietary supplement. CONCLUSIONS: The previous findings illustrated that DI-nano-ESI-MS/MS approach can emerge as a powerful, high throughput, and promising analytical tool for screening and accurate detection of various pharmaceuticals and ingredient in dietary supplements as well as biological fluids.

Vancomycin-associated retinal hemorrhages in pediatric age group: A case report.

PURPOSE: To report a case of possible post-lensectomy vancomycin-induced retinal hemorrhages in a 9-month-old infant to raise awareness of this rare postoperative complication in the pediatric age group. OBSERVATION: A retinal vascular occlusion-like findings were noted bilaterally after sequential uneventful parsplicata lensectomy in a 9-month-old infant during the very early postoperative follow-up (1-2 days). The case was recorded with no remarkable intraoperative events and received intraoperative vancomycin 20µg/ml in irrigating solution (a routine endophthalmitis prophylactic protocol). CONCLUSIONS AND IMPORTANCE: Vancomycin-associated hemorrhagic occlusive retinal vasculitis (HORV) is a rare reported postoperative complication of intraocular prophylactic vancomycin injection. Although all documented cases were reported in elderly patients aged 50 years and above, all presented with almost common findings of occlusive retinal vasculitis. To the best of author's knowledge, this is the first reported case of presumed HORV in pediatric age group. The author finds this of utmost importance to demonstrate the case to expand awareness of this possible complication in the pediatric age group as well.

Comparative study between human mesenchymal stem cells and etanercept as immunomodulatory agents in rat model of rheumatoid arthritis.

To compare human adipose tissue mesenchymal stem cells (AT-MSCs) and etanercept as immunomodulatory agents for collagen-induced arthritis (CIA). CIA was induced by rats' immunization with collagen type II (CII) in complete Freund's adjuvant in days 0 and 7. Before the onset of CIA, prevention group received five doses of AT-MSCS intraperitoneally. After establishment of arthritis, rats received either five doses of AT-MSCs or phosphate-buffered saline (PBS) intraperitoneally or six doses of etanercept subcutaneously. Clinical and histopathological evaluation were performed in all groups; serum levels of tumor necrosis factor-α (TNF-α), interleukin-10 (IL-10), and anti-collagen II were assessed by enzyme-linked immunosorbent assay (ELISA). A total percent of autoreactive T and regulatory T (Treg) cells were quantified using spleen immune histochemical analysis. AT-MSCs were able to delay the onset of CIA, suppress the ongoing clinical and histopathological signs, decrease serum levels of TNF-α and anti-collagen type II, and downregulate the autoreactive T cells as etanercept. AT-MSCs were more potent in Treg cells upregulation, producing high serum levels of IL10. AT-MSCs might have a therapeutic effect in CIA via their potency in immune cell education, representing an effective new promising approach in rheumatoid arthritis in human.

Integration of Solid-Phase Extraction and Reversed-Phase Chromatography in Single Protein-Coated Columns for Direct Injection of Bupivacaine in Human Serum.

A rapid, reliable and precise integrated solid-phase extraction (SPE) and reversed-phase liquid chromatography method was developed and validated to determine bupivacaine in human serum using single protein-coated analytical columns. The protein-coated columns were packed with four different sorbents: TSK-ODS, LiChrosorb RP-8, LiChrosorb RP-2 and µ-Bondapak CN-bonded silica. The method involved direct injection of serum sample onto the columns for trapping of the analyte, clean-up from weakly retained serum endogenous components, as well as the final separation. The protein-coated columns operated in two different chromatographic modes. Serum proteins were extracted and cleaned up by SPE, whereas the final separation of bupivacaine was based on reversed-phase chromatography. The protein-coated TSK-ODS column resulted in more accurate peak integration and more reproducible results. A linear relationship between the concentrations of drug and peak areas was confirmed in the range of 100-2000 ng/mL. Detection and quantification limits were 24.85 and 85.36 ng/mL, respectively. The average recovery for bupivacaine ranged from 96.48% to 98.81%. The present methodology was successfully applied, with a high degree of confidence, to analyze clinical samples obtained from patient receiving 0.5% bupivacaine therapy.

Direct nano-electrospray ionization tandem mass spectrometry for the quantification and identification of metronidazole in its dosage form and human urine.

A rapid, sensitive and direct nano-electrospray ionization-tandem mass spectrometry (NS-ESI-MS/MS) method, using an offline nanospray (NS) capillary, has been developed and validated for the analysis of metronidazole (MTZ). A mixture of 2 µl MTZ sample solution prepared in an ionization solvent consisting of methanol : water : formic acid in a ratio of 80 : 20 : 0.3, together with 2 µl of an internal standard (IS), 1,3,6-polytyrosine, is loaded into the back of the NS capillary. The NS capillary was fitted into the ion source at a distance of 3 mm between the NS tip and MS orifice. The sample is then analysed and acquired a sustainable signal that allowed for data compilation across various data points for MTZ identification and quantification. The quantification relied on the ratio of the [M + H]+ peaks of MTZ and IS with m/z values of 172.0717 and 182.0812, respectively, while the identification relied on the MS/MS of the precursor ions [M + H]+ of both compounds and their fragments at 128.05 for MTZ and 165.1 and 136.07 for the IS. The NS-ESI-MS/MS method was accurate and precise for the quantification of MTZ over the concentration range from 2.5 to 25 000 ng ml-1. The applicability of the method was confirmed by MTZ analysis in its pharmaceutical dosage form and detection of the analyte in clinical human urine samples without any sample treatment procedure.

The Effect of Strabismus Muscle Surgery on Corneal Biomechanics.

PURPOSE: Studying the early effect of different extraocular muscle (EOM) surgeries on corneal biomechanics. SUBJECTS AND METHODS: This is a prospective, nonrandomized, interventional study, in which 42 eyes of 29 candidates for EOM surgery for strabismus correction at Cairo university hospitals, aged 14-37 years, were recruited. All participants had measuring of the visual acuity, refraction (spherical equivalent (SE)), assessment of the EOM motility and muscle balance, sensory evaluation, fundus examination, and assessing the ocular biomechanics using the Ocular response analyzer (ORA, Reichert, INC., Depew, NY) noting the corneal hysteresis (CH) and corneal resistance factor (CRF) preoperatively. Same patients were reassessed using ORA 4 weeks postoperatively following a different standard EOM surgery (recti weakening/strengthening and inferior oblique weakening either (graded recession) according to the surgical indication, and ∆CH and ∆CRF were calculated, each is the preoperative - the postoperative value. RESULTS: ∆CH and ∆CRF = -0.78 ± 1.56 and -0.72 ± 2.15, respectively, and a highly significant difference was found between each of the pre- and postoperative CH and CRF (p < 0.001). 18 eyes had single EOM surgery, while 24 had multiple (2 or 3) EOM surgery; ∆CH in the single group = -1.28 ± 1.5, and ∆CH in the multiple group = 0.4 ± 1.49 (p=0.07). 23 eyes had EOM weakening surgery, while 18 had combined weakening and strengthening EOM surgery: ∆CH in the weakening group = -1.24 ± 1.77 and ∆CH in combined group = -0.26 ± 1.07 (p=0.04). A nonsignificant difference was found for ∆CRF (p=0.53). CONCLUSION: A different EOM surgery has an early tendency for increase of the postoperative CH specially for muscle weakening procedures (recti recession/inferior oblique muscle weakening).

Endoscopic-Assisted Scleral Fixated IOL in the Management of Secondary Aphakia in Children.

Purpose. To evaluate the short-term postoperative outcomes in endoscopic-assisted sclera fixation intraocular lens (IOL) for the management of secondary aphakia in children. Methods. This is a prospective study, whereas 40 aphakic eyes with absence of a good capsular support were implanted by endoscopy-assisted sclera fixation technique. Results. No major intraoperative complications were recorded. All cases were followed up for 6 months. Only transient ocular hypertension occurred in 10 (25%) eyes. Lens decentration and/or tilting were clinically detected in 2 eyes (5%). Ultrasonic biomicroscopic (UBM) examination revealed lens tilting in 2 (5%) of the operated eyes, despite the proper haptics positioning in the ciliary sulcus. Postoperative vitreous hemorrhage was reported in 5 eyes (12.5%) in the early postoperative period and retinal detachment in one eye. A postoperative refractive astigmatism ranging from 0.75 D to 3.75 D (mean 1.7 D ± 0.79) was recorded, as compared to mean preoperative values of 2.00 D, with no statistically significant differences being recorded (p ≥ 0.05). An improvement of BCVA, 1-2 lines on Snellen chart at the end of the follow-up period, was detected in 23 eyes (57.5%) with a mean of 0.6 ± 0.08 SD, as compared to a preoperative mean values of 0.5 ± 0.07 SD (p ≥ 0.05). Conclusion. Using an endoscope for transscleral suturing of intraocular lenses in aphakic pediatric eyes might be considered as being an effective technique that can reduce surgical complications, especially postoperative lens decentration.

A Controlled Study on the Correlation between Tear Film Volume and Tear Film Stability in Diabetic Patients.

Purpose. To assess the tear film quantity and correlate it with the quality and stability of the tear film in diabetics and compare them to age matched controls. Introduction. Diabetes affects tear film parameters in multiple ways. Poor metabolic control and neuropathy are postulated factors. To further understand how diabetes affects tear film parameters this study was conducted. Subjects and Methods. Tear meniscus height was measured by anterior segment OCT, along with tear thinning time, a subtype of noninvasive tear break-up time, and blinking rate per minute which were all recorded for 22 diabetic patients. Correlations between these tear film parameters were studied and then compared to 16 age matched controls. Results. A statistically significant difference was found in blinking rate between the diabetic and the control group (P = 0.002), with higher blinking rate among diabetics. All tear film parameters were negatively correlated with duration of diabetes. A positive correlation was found between tear film volume and stability. Conclusion. Diabetes affects the tear film in various ways. Diabetics should be examined for dry eye signs even in absence of symptoms which may be masked by associated neuropathy. Duration of diabetes has an impact on tear film status.

The Effectiveness of Intraocular Methotrexate in the Treatment of Posterior Uveitis in Behçet's Disease Patients Compared to Retrobulbar Steroids Injection.

Aim of Work. To evaluate the efficacy of intravitreal methotrexate (MTX) compared to retrobulbar triamcinolone acetonide (TAA), in controlling posterior segment involvement and inducing remissions among Behçet's disease (BD) patients. Study Design. This is a cross-sectional nonrandomized comparative study. Patients and Methods. 31 adult BD male patients with a mean disease duration of 5.45 years who presented with bilateral posterior segment involvement were included. Each patient received intravitreal injection of 400 µg/0.1 mL (MTX) for the right eye (Group A) and 1 mL of retrobulbar 40 mg/mL TAA for the left eye (Group B). Results. 90% of eyes showed complete improvement of anterior chamber reaction, whereas an improvement in vitreous activity in 77% with no significant differences between both groups (p ≤ 0.1). BCVA improved in 77.4% eyes (Group A) compared to 87.1% (Group B) (p ≤ 0.4). Relapses were noted in 11 eyes (35.5%), in group A, with the mean duration of remission being 19.1 weeks ± 2.13 compared to 7.35 ± 2.8 in 20 eyes (64.5%) in group B (p ≤ 0.1). Conclusion. No statistical differences were found between both treatment modalities; however, based on clinical observations, intravitreal MTX may ensure better control of inflammatory reaction and may encourage longer remission as compared to retrobulbar TAA in BD patients.