A pilot study of the depression, anxiety and stress in Greek military personnel during the first year of the COVID-19 pandemic.

INTRODUCTION: The ongoing SARS-CoV-2 pandemic is a global health crisis which poses many psychological research challenges. The objective of this study was to evaluate the responsiveness and validity of depression, anxiety and stress in a representative cohort of Hellenic military personnel using the short version of the Greek military version of the Depression-Anxiety-Stress Scales-21 (DASS-21) questionnaire. METHODS: A total of 158 participants were voluntarily surveyed using an electronically developed structured questionnaire. The anonymous e-questionnaire included a social demographic section and the DASS-21 scale section. It was distributed in September 2020 to the military personnel of the Hellenic Tactical Air Force Units using an intranet network during government restrictive measures due to the COVID-19 crisis. RESULTS: Our DASS-21 survey indicated a positive outcome at the psychometric level of our military sample population. Age, sex and systemic medications were statistically correlated with anxiety. Inter-correlations between the DASS-21 statements showed that individuals with low-level depression also experienced some degree of stress. Factor analysis indicated the reliability and validity of the questionnaire. CONCLUSIONS: The low levels of depression and stress among our military sample population demonstrate the importance of periodic monitoring of the psychometric items of the DASS-21 subscales to design and implement psychological prevention strategies, especially during the ongoing and future healthcare crises.

Serum microRNA array analysis identifies miR-140-3p, miR-33b-3p and miR-671-3p as potential osteoarthritis biomarkers involved in metabolic processes.

BACKGROUND: MicroRNAs (miRNAs) in circulation have emerged as promising biomarkers. In this study, we aimed to identify a circulating miRNA signature for osteoarthritis (OA) patients and in combination with bioinformatics analysis to evaluate the utility of selected differentially expressed miRNAs in the serum as potential OA biomarkers. METHODS: Serum samples were collected from 12 primary OA patients, and 12 healthy individuals were screened using the Agilent Human miRNA Microarray platform interrogating 2549 miRNAs. Receiver Operating Characteristic (ROC) curves were constructed to evaluate the diagnostic performance of the deregulated miRNAs. Expression levels of selected miRNAs were validated by quantitative real-time PCR (qRT-PCR) in all serum and in articular cartilage samples from OA patients (n = 12) and healthy individuals (n = 7). Bioinformatics analysis was used to investigate the involved pathways and target genes for the above miRNAs. RESULTS: We identified 279 differentially expressed miRNAs in the serum of OA patients compared to controls. Two hundred and five miRNAs (73.5%) were upregulated and 74 (26.5%) downregulated. ROC analysis revealed that 77 miRNAs had area under the curve (AUC) > 0.8 and p < 0.05. Bioinformatics analysis in the 77 miRNAs revealed that their target genes were involved in multiple signaling pathways associated with OA, among which FoxO, mTOR, Wnt, pI3K/akt, TGF-ß signaling pathways, ECM-receptor interaction, and fatty acid biosynthesis. qRT-PCR validation in seven selected out of the 77 miRNAs revealed 3 significantly downregulated miRNAs (hsa-miR-33b-3p, hsa-miR-671-3p, and hsa-miR-140-3p) in the serum of OA patients, which were in silico predicted to be enriched in pathways involved in metabolic processes. Target-gene analysis of hsa-miR-140-3p, hsa-miR-33b-3p, and hsa-miR-671-3p revealed that InsR and IGFR1 were common targets of all three miRNAs, highlighting their involvement in regulation of metabolic processes that contribute to OA pathology. Hsa-miR-140-3p and hsa-miR-671-3p expression levels were consistently downregulated in articular cartilage of OA patients compared to healthy individuals. CONCLUSIONS: A serum miRNA signature was established for the first time using high density resolution miR-arrays in OA patients. We identified a three-miRNA signature, hsa-miR-140-3p, hsa-miR-671-3p, and hsa-miR-33b-3p, in the serum of OA patients, predicted to regulate metabolic processes, which could serve as a potential biomarker for the evaluation of OA risk and progression.

miR-15a and miR-24-1 as putative prognostic microRNA signatures for pediatric pilocytic astrocytomas and ependymomas.

In the current setting, we attempted to verify and validate miRNA candidates relevant to pediatric primary brain tumor progression and outcome, in order to provide data regarding the identification of novel prognostic biomarkers. Overall, 26 resected brain tumors were studied from children diagnosed with pilocytic astrocytomas (PAs) (n = 19) and ependymomas (EPs) (n = 7). As controls, deceased children who underwent autopsy and were not present with any brain malignancy were used. The experimental approach included microarrays covering 1211 miRNAs. Quantitative real-time polymerase chain reaction (qRT-PCR) was performed to validate the expression profiles of miR-15a and miR-24-1. The multiparameter analyses were performed with MATLAB. Matching differentially expressed miRNAs were detected in both PAs and EPs, following distinct comparisons with the control cohort; however, in several cases, they exhibited tissue-specific expression profiles. On correlations between miRNA expression and EP progression or outcome, miR-15a and miR-24-1 were found upregulated in EP relapsed and EP deceased cases when compared to EP clinical remission cases and EP survivors, respectively. Taken together, following several distinct associations between miRNA expression and diverse clinical parameters, the current study repeatedly highlighted miR-15a and miR-24-1 as candidate oncogenic molecules associated with inferior prognosis in children diagnosed with ependymoma.

MicroRNA expression profiles in pediatric dysembryoplastic neuroepithelial tumors.

Among noncoding RNAs, microRNAs (miRNAs) have been most extensively studied, and their biology has repeatedly been proven critical for central nervous system pathological conditions. The diagnostic value of several miRNAs was appraised in pediatric dysembryoplastic neuroepithelial tumors (DNETs) using miRNA microarrays and receiving operating characteristic curves analyses. Overall, five pediatric DNETs were studied. As controls, 17 samples were used: the FirstChoice Human Brain Reference RNA and 16 samples from deceased children who underwent autopsy and were not present with any brain malignancy. The miRNA extraction was carried out using the mirVANA miRNA Isolation Kit, while the experimental approach included miRNA microarrays covering 1211 miRNAs. Quantitative real-time polymerase chain reaction was performed to validate the expression profiles of miR-1909* and miR-3138 in all samples initially screened with miRNA microarrays. Our findings indicated that miR-3138 might act as a tumor suppressor gene when down-regulated and miR-1909* as a putative oncogenic molecule when up-regulated in pediatric DNETs compared to the control cohort. Subsequently, both miRNA signatures might serve as putative diagnostic biomarkers for pediatric DNETs.

Optic nerve head neuroretinal rim blood flow differences in monkeys with laser-induced glaucoma.

INTRODUCTION: The blood flow of the neuroretinal rim (NRR) of the optic nerve head (ONH) of the rhesus monkey with laser-induced glaucoma was examined. METHODS: Argon laser photocoagulation of the trabecular meshwork to induce elevated intraocular pressure (IOP) was performed in one eye of nine normal male rhesus monkeys. The nasal and temporal NRR of the monkey ONH were examined by the Heidelberg retina tomograph/flowmeter (HRT/HRF) under neuromuscular blockade. A mixed effect analysis of variance was used to determine significant differences between eyes and between locations in the eyes. RESULTS: The average IOP in the hypertensive glaucoma and normal eyes was 34.8 +/- 7.2 and 16.0 +/- 1.9 mmHg, respectively. The HRT determined average overall cup to disc (C/D) area ratio in the glaucoma and normal eyes, which was 0.49 +/- 0.28 and 0.22 +/- 0.16, respectively. The mean temporal NRR HRF flow in the hypertensive eyes was significantly greater than in the normotensive eyes (P < 0.0001), than in the nasal NRR of the hypertensive eyes (P < 0.0001) and than in the nasal NRR of the normotensive eyes (P < 0.01). The mean nasal NRR HRF flow in the hypertensive eyes was significantly less than in the nasal NRR of the normotensive eyes (P < 0.01). There was no statistical difference between the mean HRF flow of the temporal and nasal NRR of the normotensive eyes. The elevated IOP positively influenced the flow values in the hypertensive eye (r = 0.724). CONCLUSIONS: The capillary microcirculation of the temporal NRR of the rhesus monkey ONH with laser-induced glaucoma has significantly increased blood flow, and the nasal NRR significantly reduced blood flow compared to blood flow in the NRR of normal normotensive monkey eyes.

Time-specific intraocular pressure curves in Rhesus macaques (Macaca mulatta) with laser-induced ocular hypertension.

OBJECTIVE: To detect and categorize time-specific variations in daytime intraocular pressure (IOP) found in Rhesus monkeys with laser-induced ocular hypertension. PROCEDURES: Ten male monkeys with argon laser-induced ocular hypertension in one eye were anesthetized with ketamine hydrochloride, and the IOP measured in both eyes at 7 a.m., 7.30 a.m., and then hourly until 1 p.m. with a Tonopen trade mark XL applanation tonometer. Intraocular pressure time profiles for both eyes in each animal were developed. The means +/- SD of the IOPs for both eyes were calculated for the whole 6-h study period, and the values compared statistically. The difference between the lasered eye mean IOP standard deviation and the normal eye mean IOP standard deviation for each animal during the 6-h follow-up was also calculated and compared. RESULTS: Mean IOP (+/- SD) in the glaucoma and normal eyes for the 10 animals during the 6-h study was 32.6 +/- 2.5 and 14.9 +/- 2.5 mmHg, respectively. The IOP was significantly higher in the experimental eye than in the normal eye (P = 0.0008). The mean IOP in the lasered eye did not significantly change during the study period, whereas a slight but significant increase in IOP of the normal eye over the study period was recorded (P = 0.003). The variance in IOP in the hypertensive eyes was considerably greater than that in the untreated control eyes. From 7 a.m. to 1 p.m. the IOP declined in five eyes and increased in the other five eyes with laser-induced ocular hypertension. CONCLUSIONS: The time-specific IOP variation pattern in the daytime in the laser treated eyes is significantly greater than the variation in the normotensive eyes. This shows that in order to detect statistical differences between IOP variations induced by an IOP-reducing drug, and the exaggerated spontaneous IOP variations present in the laser-induced hypertensive eye, sufficient animals should be included in any study. Understanding the time-specific IOP variation present in a group of monkeys with laser-induced ocular hypertension is essential prior to using the model for the evaluation of IOP-reducing drugs.

Mechanisms of action of unoprostone on trabecular meshwork contractility.

PURPOSE: This study was performed to clarify the possible mechanism behind the ocular hypotensive effect of unoprostone isopropyl (Rescula; Novartis Ophthalmics AG, Basel, Switzerland), a new docosanoid that has been shown to reduce intraocular pressure (IOP) in patients with ocular hypertension or primary open-angle glaucoma. To gain insight into the possible mode of action, the effects of unoprostone on ciliary muscle (CM) and trabecular meshwork (TM) contractility, intracellular calcium levels, and membrane channels were investigated. METHODS: The effects of unoprostone (M1 metabolite = free acid, 10(-5) M) and endothelin (ET)-1 (10(-9) M) on bovine TM (BTM) and ciliary muscle (CM) strips were investigated, by using a custom-made force-length transducer system. The effects of unoprostone and ET-1 (5 x 10(-8) M) on intracellular Ca(2+) mobilization in cultured human TM (HTM) were measured using fura-2AM as a fluorescent probe. Patch-clamp experiments were performed on HTM and BTM cells to investigate the unoprostone-dependent modulation of membrane currents. RESULTS: In isolated TM and CM strips, unoprostone almost completely inhibited ET-induced contractions (TM: 2.9% +/- 4.3% vs. 19.6% +/- 5.7%, P < 0.05, n = 6; CM: 1.4% +/- 1.6% vs. 30.1% +/- 5.3%, P < 0.01, n = 6; 100% = maximal carbachol-induced (10(-6) M) contraction). However, neither carbachol-induced contraction nor baseline tension was affected by unoprostone. Furthermore, unoprostone had no effect on baseline intracellular calcium levels (baseline: 126 +/- 45 nM versus unoprostone: 132 +/- 42 nM, n = 8) in HTM cells. The endothelin-induced increase (679 +/- 102 nM), however, was almost completely (P < 0.01) blocked by unoprostone (178 +/- 40 nM). In patch-clamp recordings, unoprostone could be shown to double the amplitude of outward current (HTM: 200% +/- 33%; n = 6; BTM: 179% +/- 20%; n = 8). This effect was blocked by the specific inhibitor of maxi-K channels, iberiotoxin. CONCLUSIONS: This study presents evidence for direct interaction of unoprostone with the contractility of the TM and CM. This compound may lower IOP by affecting aqueous outflow, most probably conventional outflow pathways (i.e., TM) through inhibition of ET-dependent mechanisms. In addition, unoprostone interacts with the maxi-K channel. Although primarily Ca(2+)-sensitive signal-transduction pathways seem to be involved, effects of unoprostone on Ca(2+)-independent pathways and uveoscleral outflow cannot be excluded.

Hypoglycemia induces general neuronal death, whereas hypoxia and glutamate transport blockade lead to selective retinal ganglion cell death in vitro.

PURPOSE: To examine the impact of experimental ischemia and interruption of glutamate transport on retinal neuronal cell, especially retinal ganglion cell (RGC), survival in vitro. METHODS: Cell cultures were prepared from adult pig retinas and maintained under different experimental conditions of increasing hypoglycemia, environmental hypoxia (delayed postmortem period or atmospheric PO2 <2%), or chemical hypoxia (potassium cyanide), or in the presence of glutamate transporter blockers L-trans-pyrrolidine-2,4-dicarboxylic acid (tPDC) and L(-)-threo-3-hydroxyaspartic acid (THA), or the glutamine synthetase inhibitor methionine sulfoximine (MS). After 48 hours, cells were returned to standard culture conditions and allowed to develop for 5 days, when they were fixed and immunostained with different retinal neuronal phenotypic markers. RESULTS: Control normoxic cultures contained large numbers of immunocytochemically identified photoreceptors (PRs), bipolar cells (BCs), amacrine cells (ACs), and RGCs after 7 days in vitro. A 24-hour postmortem delay before culture led to significant reductions in all types (40%-70%), proportionately greater in ACs and RGCs. Lowering of sugar levels also led to increased losses in all cell types, whereas potassium cyanide treatment deleteriously affected only ACs and RGCs. Ambient hypoxia led to consistent reductions only in the number of RGCs, which were exacerbated by addition of high concentrations of glutamate. Inclusion of glutamate receptor antagonists had a partial protective effect against RGC loss. Treatment with tPDC and THA also led to selective RGC death, but MS had no effect on any cells. CONCLUSIONS: Different components of the ischemic pathologic process (hypoxia, hypoglycemia, glutamate transport failure) lead to distinctly different patterns of neuronal loss in adult retina in vitro. RGCs are especially vulnerable, corresponding to their in vivo susceptibility. These data may suggest neuroprotective strategies for limiting retinal damage during ischemia.

Comparison of the vasoactive effects of the docosanoid unoprostone and selected prostanoids on isolated perfused retinal arterioles.

PURPOSE: To compare the vasoactive properties of the docosanoid unoprostone, its free acid, and different members of the prostanoid family on isolated perfused pig retinal arterioles to assess their potential to modulate retinal blood flow. METHODS: Segments of porcine retinal arterioles were dissected, cannulated, and perfused, and their diameter monitored during either intraluminal or extraluminal application of increasing doses (10(-10)-10(-4) M) of either the docosanoid unoprostone isopropyl and its free acid or of selected prostanoids: prostaglandin (PG) F(2alpha) and thromboxane A(2) analogue (U46619). Studies were performed on arterioles in their uncontracted state, and also during precontraction with endothelin-1 (10(-9) M). The significance of any induced change in vessel diameter was assessed in relation to the initial vessel diameter or, in the case of endothelin-1 administration, to the contracted diameter with endothelin-1 alone. RESULTS: In normal-tone arterioles without endothelin-1 contraction, PGF(2alpha) and U46619 both produced a potent dose-dependent contraction, but neither unoprostone isopropyl nor unoprostone free acid had a significant vasoactive effect. In endothelin-1-contracted arterioles, U46619 produced further contraction, PGF(2alpha) produced a slight vasodilatation, and unoprostone isopropyl and its free acid produced a pronounced dilatation. CONCLUSIONS: Of the agents tested, unoprostone isopropyl and its free acid were the most potent vasodilators of endothelin-1-contracted pig retinal arterioles. Members of the prostanoid family demonstrated a different effect on the diameter of isolated retinal arterioles compared with the docosanoids. The potential therefore exists for the docosanoid unoprostone to have a beneficial effect on retinal blood flow in addition to any reduction in intraocular pressure.

Selective excitotoxic degeneration of adult pig retinal ganglion cells in vitro.

PURPOSE: Excitotoxicity is proposed to play a prominent role in retinal ganglion cell (RGC) death ensuing from diseases such as glaucoma and ischemia, but cell culture studies have used tissue from newborn rodents, yielding conflicting data that implicate either N-methyl D-aspartate (NMDA) or non-NMDA glutamate (Glu) receptor-mediated pathways. Excitotoxic RGC death was examined in vitro in this study, using adult pigs, a large-animal model for human retina. METHODS: Adult pig retina (and for comparative purposes young and adult rat retina) were dissociated and maintained in monolayer culture. Medium was supplemented with Glu or pharmacologic agonists or antagonists, and surviving RGCs and other retinal neurons were quantified using specific immunolabeling methods. Electrophysiological responses to externally applied Glu of RGCs in culture were recorded using whole-cell patch-clamp techniques. RESULTS: Application of Glu led to selective, dose-dependent losses in large RGCs (maximal 37% decrease at 1 mM; median effective dose [ED50], approximately 80 microM) and neurite damage in surviving RGCs. Application of Glu agonists and Glu receptor subclass antagonists showed that large RGC death was mediated through both NMDA and non-NMDA receptor pathways. Small RGCs, amacrine cells, and all other retinal neurons were resistant to Glu-induced death. By comparison, rat retinal cultures displayed heightened RGC vulnerability to Glu, mediated exclusively by non-NMDA receptor-mediated pathways. Amacrine cells were unaffected by NMDA but were very sensitive to kainate application (>90% loss). Other retinal neurons were unaffected by any treatment. CONCLUSIONS: The molecular pathways underlying excitotoxic RGC death in vitro (non-NMDA or NMDA-preferring Glu receptors) vary among species and developmental stages. The selective elimination of adult pig large RGCs by NMDA receptor-mediated pathways more closely resembles human and animal glaucoma in vivo than other published culture models, providing a simplified experimental system for investigating the pharmacologic and toxicologic bases of glaucoma-like neuronal death.

The effect of 0.12% unoprostone isopropyl (rescula) on intraocular pressure in normotensive dogs.

Rescula (0.12% unoprostone isopropyl) is the first docosanoid compound approved for treatment of glaucoma in humans. It is commercially available in Japan, and is undergoing clinical testing elsewhere. The aim of this study was to evaluate the effect of Rescula on intraocular pressure (IOP) in normotensive dogs. After establishing a baseline diurnal IOP curve, six dogs were unilaterally treated with Rescula while the contralateral eye was treated with a placebo. Applanation tonometry was performed in both eyes, and pupil size was evaluated, 30 min after treatment, and at 1-hr intervals for the next 9 hr. Rescula caused a significant (p=0.014) and long-lasting decrease in IOP, from 20.49+/-2.02 mm Hg in control eyes to 15.49+/-0.69 mm Hg in treated eyes. These results suggest that Rescula is potentially efficacious in treatment of canine glaucoma.

Flow ratios to express results obtained with the human in vivo 'perfused forearm technique'.

AIMS: To determine the accuracy of forearm blood flow (FBF) ratio (flow in infused arm/flow in control arm) to detect unilateral increases in forearm blood flow. METHODS: In nine healthy male volunteers, we measured the effect of infusion of saline into the brachial artery at a rate of 2 ml/100 ml forearm min-1 on FBF ratio during control, mental arithmetic (MAR) and lower body negative pressure (LBNP) at -40 mmHg. RESULTS: Saline infusion increased FBF ratio from baseline by 115.9+/-17.4, 82.0+/-19.0 and 159.6+/-53.3% for control, MAR and LBNP, respectively (P<0.05 for MAR vs control). CONCLUSIONS: FBF ratio may underestimate unilateral increases in forearm blood flow during simultaneous mental arousal.

Ocular blood flow in experimental glaucoma: a study in cynomolgus monkeys.

Experimental glaucoma was induced in 1 eye of 6 cynomolgus monkeys by laser treatment of the trabecular meshwork. In 5 of the 6 monkeys the increased intraocular pressure (IOP) caused marked glaucomatous damage in the experimental eye. Ocular blood flow was determined with labeled microspheres 4 years after the laser treatment. IOP was regulated with an external reservoir. With the same perfusion pressure in both eyes no statistically significant difference was observed between the 2 eyes for total ocular blood flow or for blood flow through any of the ocular tissues. Total ocular blood flow was 343.5 +/- 61.4 mg/min (mean +/- SEM) in the control eye and 385.3 +/- 107.7 mg/min in the experimental eye.

Isolation of a novel RFamide peptide from the midgut of the American cockroach, Periplaneta americana.

In insects (FM)RFamide-immunoreactive endocrine cells are ubiquitously present in the midgut, but the identity of the peptide(s) produced by these cells is unknown. The major RFamide-immunoreactive peptide from the midgut of the American cockroach, Periplaneta americana, was isolated and identified as Ala-Asn-Arg-Ser-Leu-Arg-Leu-Arg-Pheamide. This is a novel member of an arthropod peptide family, previously known only from mosquitoes and horseshoe crabs. Its abundance in the midgut suggests that it plays an important function in digestion.

Hemodynamic parameter estimation from ocular fluorescein angiograms.

BACKGROUND: A method is proposed for parameterizing choroidal blood flow from fluorescein angiograms. METHODS: After digitizing and aligning the angiographic sequence, the intensity build-up curves of fluorescence are analysed per pixel (approx. 10 microns in fundo). Two models are compared. A one-compartment model predicts an exponential build-up curve, from which the following parameters are estimated: maximum fluorescence, dye appearance time and local perfusion rate (reciprocal of the time constant of the exponential). To account for the contribution of the systemic circulation to the shape of the build-up curve, a two-compartment model is used which predicts a bi-exponential curve. RESULTS: Introduction of the second (systemic) compartment resulted in a significant improvement of fit in 37 of 48 patients studied. The rate constants of the systemic compartment found were mainly in the range of 0.30-1.00 s-1. CONCLUSION: For the individual patient, the local perfusion rates may vary strongly, with lower perfusion rates possibly being of prognostic value for ocular diseases such as glaucoma or diabetic retinopathy.

[Slowly reversible central scotoma: iatrogenic effect of hyperbaric oxygenation in the treatment of multiple sclerosis]. / Le scotome central lentement réversible: un effet iatrogène de l'oxygénothérapie hyperbare dans le traitement de la sclérose en plaques.

Considerable enthusiasm has been raised in the past about the use of Hyperbaric Oxygen (HBO) in various diseases, usually otherwise untreatable. Recently, special attention has been drawn on its hypothetical beneficial effects on multiple sclerosis (MS). We have witnessed a rare, though known, side-effect of HBO on a patient suffering from MS. She developed an acute, bilateral, centro-caecal scotoma, from which she slowly recovered several days after. The forementioned case led us to a review of the literature concerning: Various attempts to employ HBO in ophthalmology Side-effects of oxygen on eye and vision Possible mechanisms of ocular toxicity of oxygen. It appears from this review that we should be extremely cautious about using HBO on MS patients, particularly able to develop such side-effects.