ACUTE KERATOUVEITIS WITH CORNEAL GRAFT MELTING AS A LATE COMPLICATION OF MILD CHEMICAL BURN POTENTIALLY ASSOCIATED WITH COVID-19 INFECTION: A CASE REPORT.

Chemical burns are sight-threatening injuries that require immediate management. The main goal of the initial treatment is prompt and copious irrigation to neutralize and eliminate the chemical, followed by various therapeutic options (conservative and surgical) according to the individual patient's postinjury findings. PURPOSE: In this case report, we describe the course and treatment of a patient with chemical alkali burns of both eyes. This study reports the outcomes of an ocular alkali burn patient who became infected with COVID-19 and where severe keratouveitis with corneal graft melting and hypopyon occurred, ultimately resulting in evisceration of the eye. OBSERVATIONS: We report the case of a 35-year-old woman, after an alkali burn of both eyes. Complete re-epithelialization of both corneas occurred within three weeks. Due to this observation, we assumed a satisfactory healing prognosis. However, corneal lysis occurred in the left eye over time. The condition was managed, but subsequently severe keratouveitis with hypopyon and corneal lysis occurred in the left eye, apparently in connection with the COVID-19 infection, which ultimately led to the evisceration of the affected eye. CONCLUSIONS AND IMPORTANCE: In the case report, we describe the management of a mild chemical alkali burn of the right eye and a complicated moderate burn of the left eye with an alkaline chemical. Both eyes were completely healed within three weeks. However, the findings on the left eye became complicated, which ultimately led to the evisceration of the affected eye. The disease of COVID-19 could have contributed to the deterioration of the findings on the cornea, or it could have just been a coincidence of two diseases, with the keratouveitis having a serious course due to the previous difficult course of healing and many complications after the chemical burn.

Leg raise increases pressure in lower and upper esophageal sphincter among patients with gastroesophageal reflux disease.

The purpose of this study was to determine the relation between posturally increased intra-abdominal pressure and lower/upper esophageal sphincter pressure changes in patients with gastroesophageal reflux disease. We used high resolution manometry to measure pressure changes in lower and upper esophageal sphincter during bilateral leg rise. We also examined whether the rate of lower and upper esophageal sphincter pressure would increase during leg raise differentially in individuals with versus without normal resting pressure. Fifty eight patients with gastroesophageal reflux disease participated in the study. High resolution manometry was performed in relaxed supine position, then lower and upper esophageal sphincter pressure was measured. Finally, the subjects were instructed to keep their legs lifted while performing 90-degree flexion at the hips and knees and the pressure was measured again. Paired t-test and independent samples t-test were used. There was a significant increase in both lower (P < 0.001) and upper esophageal sphincter pressure (P = 0.034) during leg raise compared to the initial resting position. Individuals with initially higher pressure in lower esophageal sphincter (>10 mmHg) exhibited a greater pressure increase during leg raise than those with initially lower pressure (pressure ≤10 mmHg; P = 0.002). Similarly individuals with higher resting upper esophageal sphincter pressure (>44 mmHg) showed a greater pressure increase during leg raise than those with lower resting pressure (≤44 mmHg; P < 0.001). The results illustrate the influence of postural leg activities on intraesophageal pressure in patients with gastroesophageal reflux disease, indicating by means of high resolution manometry that diaphragmatic postural and sphincter function are likely interrelated in this population.

[Patophysiology of diabetic retinopathy]. / Patofyziologie diabetické retinopatie.

Diabetic retinopathy (DR) is the affection of the retina in patients with diabetes mellitus (DM). The basic causative factor is prolonged hyperglycaemia. DR is microangiopathy, ie impairment of retinal capillaries. Pathophysiology of DR is very complex and there are involved in many factors. The first and most fundamental factor is the failure blood-retinal barrier (BRB). The major mechanism causing malfunction of BRB are advanced glycation end-products (AGE). In the failure of the inner BRB are involved losses of endothelial cells in capillaries, together with the losses of pericytes. A very important role in the failure of BRB plays too increased adhesivity of leukocytes. Further important role play also AGE and their receptor RAGE. They stimulate cascade of pathological processes damaging BRB. The second important factors in the pathophysiology of DR are vasoactive factors. The most important is vascular endothelial growth factor (VEGF), further than protein kinase C (PKC), histamine, angiotensin II, matrix metaloproteinases. The third important factor in the pathophysiology of DR is the vitreoretinal interface. There plays important role detachment of posterior vitreous, cortical vitreous, internal limiting membrane.

[Photodynamic therapy with Verteporfin in treatment of wet form ARMD - long term results]. / Lécba vlhké formy vekem podmínené makulární degenerace fotodynamickou terapií s verteporfinem - dlouhodobé výsledky.

Photodynamic therapy with Verteporfin (Visudyne - Novartis AG, Basel, Switzerland) is a method designed for treatment of the wet form of age-related macular degeneration. Effectiveness of photodynamic therapy with Verteporfin we approved on our department in group of 301 patients (114 males, 187 women) of average age 73.5 years with predominantly classic choroidal neovascular membrane in subfoveal localization in wet form of age-related macular degeneration. The follow up period in this group was from 6 to 36 month (mean 21 months). Patients with predominantly classic choroidal neovascular membrane underwent during the observation period from 1 to 5 treatments (mean 1.45 treatments). Mean best corrected visual acuity was before treatment 0.708 ± 0.24 logMAR. At the end of three-year observation period was the mean best corrected visual acuity 1.016 ± 0.36 logMAR. Best corrected visual acuity dropped during this time by 3.08 lines (15.4 letters) of ETDRS (early treatment diabetic retinopathy study) visual charts. Decrease of mean best corrected visual acuity less than 3 lines on ETDRS charts is considered as stabilisation. This goal was achieved in our group during observation period in 2/3 of patients.

[Photodynamic therapy with verteporfin in treatment of myopic neovascular choroideal membranes]. / Vyuzití fotodynamické terapie v lécbe myopických neovaskulárních membrán.

The aim of this study was to evaluate efficiency and long-term stabilization effect in patients with classic type of myopic choroidal neovascular membrane (CNV), treated with photodynamic therapy (PDT) with verteporfin (Visudyne - Novartis AG, Basel, Switzerland). We have verified the efficiency of photodynamic therapy with verteporfin in group of 51 eyes (17 men, 34 women), mean age 49,5 years with subfoveal localized predominantly classic neovascular membranes in pathologic myopia. The average follow up period was 23,7 months (± 2,3 month). Patients underwent during follow-up period 1 to 3 sessions of photodynamic therapy (PDT average number 1,25 sessions). The average best corrected visual acuity (BCVA) before treatment was 0,302 (0,65 logMAR) and the average BCVA at the end of follow up was 0,356 (0,46 log MAR). The improvement of best corrected visual acuity up to 5 letters on ETDRS (Early Treatment Diabetic Retinopathy Study) charts was observed in 23% of patients at the end of follow up.

[Ranibizumab in the ARMD wet form of age-related macular degeneration treatment - two years results obtained from the AMADEuS Registry]. / Ranibizumab v lécbe vlhké formy VPMD - dvouleté výsledky lécby z registru AMADEuS.

AIM: The aim of this study was the retrospective follow up of Age-Related Macular Degeneration (ARMD) wet form patients treated with ranibizumab during 24 months period. The data were recorded into the AMADEuS (Age-related MAcular DEgeneration in patientS in the Czech Republic) Registry and after their evaluation compared with treatment results obtained from other departments of ophthalmology collaborating in the AMADEuS project or results of some foreign studies as well. PATIENTS AND METHODS: The group consisted of patients registered since October 1, 2008 until June 11, 2012, followed up for 24 months period. There were 90 eyes of 89 patients. All patients were completely examined in the Macular ambulance of the Department of Ophthalmology in the Faculty Hospital Brno-Bohunice, Czech Republic, E.U., and consequently the ranibizumab (Lucentis, Novartis) was applied intravitreally in three initials doses one month apart. Thereafter, ranibizumab was applied "on demand". In 43.3 % of eyes the mostly classical, in 27.8 % of eyes occult, and in 28.9 % of eyes the minimally classical choroid neovascular membrane was present. The initial visual acuity was in 3.3 % of eyes in the range 15 - 30 letters of ETDRS optotypes (20/500 - 20/200), in 61.1 % of eyes in the range 31 - 60 letters (20/200 - 20/63), and the visual acuity better than 61 letters of ETDRS optotypes (better than 20/63) was in 35.6 % of eyes. RESULTS: The average initial best-corrected visual acuity (BCVA) in our group of patients was 54.2 letters of EDTRS (SD ± 14.4). At the visit at three months after the start of the treatment the BCVA was 59.6 letters of EDTRS (SD ± 15.0), at the visit after 6 months 57.3 letters of EDTRS (SD ± 14.7), after one year of the study 54.8 letters of EDTRS (SD ± 16), after 18 months of the study 53.4 letters of EDTRS (SD ± 16,8), and after 24 months of the study was the BCVA 51.7 letters of EDTRS (SD ± 16.9). The average CRT (central retinal thickness) value by means of the OCT (optic coherence tomography) examination was at the beginning of the treatment 311.4 µm (SD ± 117.9), after 3 months of treatment 233.5 µm, (SD ± 85.4), after 6 months of treatment 262.2 µm, (SD ± 102,4), after 12 months 261 µm (SD ± 88,4), after 18 months 254.9 µm (SD ± 70.0), and after 24 months 249 µm (SD ± 87.5). The average number of ranibizumab doses during the follow-up period was 5.6. After the 24 months follow-up period, the gain of 15 or more letters of EDTRS was recorded in 11.1 % of patients, the gain of 1 - 14 letters of EDTRS optotypes was recorded in 32.2 % of patients, the decrease of 14 or less letters of EDTRS optotypes was found in 21.2 % of patients, and the decrease of 15 or more letters was found in our group in 22.2 % of patients. CONCLUSION: The ARMD wet form treatment using ranibizumab is up to date the most effective available therapy. The AMADEuS registry is of great importance in the reviewing of the effectiveness of the ARMD wet form treatment.

CD73 and CD29 concurrently mediate the mechanically induced decrease of migratory capacity of mesenchymal stromal cells.

e assumption that mesenchymal stromal cell (MSC)-based-therapies are capable of augmenting physiological regeneration processes has fostered intensive basic and clinical research activities. However, to achieve sustained therapeutic success in vivo, not only the biological, but also the mechanical microenvironment of MSCs during these regeneration processes needs to be taken into account. This is especially important for e.g., bone fracture repair, since MSCs present at the fracture site undergo significant biomechanical stimulation. This study has therefore investigated cellular characteristics and the functional behaviour of MSCs in response to mechanical loading. Our results demonstrated a reduced expression of MSC surface markers CD73 (ecto-5'-nucleotidase) and CD29 (integrin ß1) after loading. On the functional level, loading led to a reduced migration of MSCs. Both effects persisted for a week after the removal of the loading stimulus. Specific inhibition of CD73/CD29 demonstrated their substrate dependent involvement in MSC migration after loading. These results were supported by scanning electron microscopy images and phalloidin staining of actin filaments displaying less cell spreading, lamellipodia formation and actin accumulations. Moreover, focal adhesion kinase and Src-family kinases were identified as candidate downstream targets of CD73/CD29 that might contribute to the mechanically induced decrease in MSC migration. These results suggest that MSC migration is controlled by CD73/CD29, which in turn are regulated by mechanical stimulation of cells. We therefore speculate that MSCs migrate into the fracture site, become mechanically entrapped, and thereby accumulate to fulfil their regenerative functions.

CTLA-4 (CD152) blockade does not cause a pro-inflammatory cytokine profile in regulatory T cells.

OBJECTIVES: The activation of T cells is closely regulated. One cell intrinsic mechanism is based on the expression of inhibitory molecules; another is mediated by regulatory T (Treg) cells. The co-regulatory molecule CTLA-4 is constitutively expressed by Treg cells and up-regulated in effector-T-cells after activation. Recently, it was described that Treg cells can display an unstable phenotype and convert into pathogenic pro-inflammatory cytokine secreting cells. Here we have analysed the role of CTLA-4 in the regulation of cytokine production by T-helper (Th) cells with a special focus on Treg cells. METHODS: Proliferation of unstimulated CTLA-4 knock-out and wild-type cells as well as their activation status and the impact of CTLA-4 blockade on proliferation of Treg and effector T cells under stimulation were analysed by flow cytometry. Furthermore, the cytokine concentrations were analysed by a multiplex suspension assay. RESULTS: CTLA-4 knock-out T cells proliferated without stimulation and displayed an activated phenotype ex vivo. Proliferation of effector but also that of Treg cells was controlled by CTLA-4. The blockade of CTLA-4 led to an increased secretion of GM-CSF, IL-1ß, IL-2, and IFN-γ by Th cells. However, the blockade of CTLA-4 in Treg cells did not cause any conversion into pathogenic pro-inflammatory T cells, since the non-cytokine secreting phenotype remained unchanged. CONCLUSIONS: These results have major implications on therapies targeting the CTLA-4-system, e.g. by CTLA4-Ig or anti-CTLA-4-antibodies, as the blockade of CTLA-4 did not unlock the stability of Treg cells.

[Epidemiology of the age-related macular degeneration]. / Epidemiologie vekem podmínené makuárni degenerace.

The age-related macular degeneration is the most common cause of severe decrease of the visual acuity in industrial countries. The epidemiological data published in last 30 years are very alarming. They document that the age-related macular degeneration is the most common cause of blindness in industrial countries in the population of patients older than 65 years. The signs of beginning disease may be diagnosed as early as in the patients older than 45 years. The disease is strictly age-related and its prevalence and incidence is markedly increasing according to the older age. The basic risk factors of the disease are smoking, untreated hypertension, atherosclerosis, cataract surgery, and appearance of the disease in the family. Among further possible risk factors we count diabetes mellitus and the light color of the iris. Equivocal is the negative influence of ultraviolet rays.

[The influence of the idiopathic macular hole (IMH) surgery with the ILM peeling and gas tamponade on the electrical function of the retina]. / Vliv operace idiopatické makulární díry s peelingem MLI a plynnou tamponádou na elektrickou funkci sítnice.

Many contemporary clinical papers establish positive influence of the pars plana vitrectomy (PPV) with the ILM (internal limiting membrane) peeling and gas tamponade in macular hole to the macular morphology. They prove diminishing or disappearing of the central scotoma and metamorphopsia and especially also improvement of the BCVA for far and near. The evaluation of the objective functional condition of the retina is still a discussed question. This paper concerns with the comparison of the electric functions of the retina before and after the IMH surgery with the ILM peeling and the gas tamponade. In the group 19 patients (8 men, 11 women), or 19 eyes with IMH were included. The average age was 69 +/- 6 years. The group consisted of patients with transparent optical media. In none of these patients was found other macular pathology than IMH. Nobody underwent other retinal surgery. The patients were examined 1 day before and 1 and 3 months after the surgery. During each control, the following examinations were performed: the Amsler grid examination, the best corrected visual acuity (BCVA) for far (EDRTS chart) and near (Jaeger optotypes), intraocular pressure measurement (non contact tonometer NIDEK NT-2000), examination of the anterior segment on the slit lamp, examination of the posterior segment biomicroscopically and by means of indirect ophthalmoscopy, examination of the photopic, pattern, and multifocal ERG (Retiscan, according to the ISCEV methodology), and OCT examination (Stratus OCT). If necessary, the ultrasound examination (Ultrascan Alcon) was performed as well. For the statistical evaluation of the ERG component values among the data files before the surgery (data file 1), 1 month after the surgery (data file 2), and 3 months after the surgery (data file 3), the non-parametric Wilcoxon pair test was used. In the photopic ERG, there was statistically significant prolongation of the latency b in data file 2 and 3 comparing to the data file 1 (p < 0.05). Comparing latency b of data file 1 to data file 2, there was found no statistical significance. Comparing other parameters of photopic ERG found no statistically significant difference among data files 1,2, and 3. In the multifocal ERG, there was found statistically significant elevation of the P1 amplitude according to the response density of given unit and the P1 amplitude in the central ring in data file 3 comparing to the data file 1 (p < 0.05). Comparison of other parameters was not statistically significant. In the paracentral ring, there was found statistically significant extension of the N1 and P1 latency in data file 3 comparing to the data file 1 (p < 0.05). Comparison of other parameters in the paracentral ring was not statistically significant. Statistically significant improvement of the retinal electric function in the central 4 degrees 3 months after the surgery, confirms the positive functional effect of the surgery to the fovea. In the fovea, the increase of the number of functional nerve cells of the outer layers of the retina occurs. On the other hand, in the parafoveolar region, as well as in the whole retina, 3 months after the surgery, statistically significant decrease of the function of the retina, meaning the time prolongation of the conduction in the outer layers of the retina, occurs. According also to our results, the peeling of the ILM in the IMH surgery remains, despite its unquestionable contribution, still a controversial technique. During the short, three months lasting, follow-up period, the functional improvement in the fovea occurred, but the functional decrease in the parafoveolar region which correlates in the large extent with area of the ILM peeling was found. The discussion about the ILM peeling indication in the earlier stages is adequate. We will further follow-up the development of the retinal electric function after the IMH surgery with ILM peeling and gas tamponade.

Stabilizing function of the diaphragm: dynamic MRI and synchronized spirometric assessment.

The aim was to describe diaphragmatic behavior during postural limb activities and examine the ventilatory and stabilizing functions of the diaphragm. Thirty healthy subjects were examined in the supine position using a dynamic MRI system assessed simultaneously with specialized spirometric readings. The diaphragmatic excursions (DEs) were measured at three diaphragmatic points in the sagittal plane; the diaphragm positions (DPs) as related to a reference horizontal baseline were determined. Measurements were taken during tidal breathing (TB) and isometric flexion of upper or lower extremities against external resistance together with TB. Mean DE in both upper and lower postural limb activities was greater compared with the TB condition (P < 0.05), with the effect greater for lower limb activities. Inspiratory DPs in the upper and lower extremity activities were lower compared with TB alone (P < 0.01). Expiratory DP was lower only for lower extremity activities (P < 0.01). DP was most affected at the apex of the crescent and crural (posterior) portion of the diaphragm. DEs correlated strongly with tidal volume (Vt) in all conditions. Changes in DEs relative to the initial value were minimal for upper and lower extremities but were related to lower values of Vt (P < 0.03). Significant involvement of the diaphragm in the limb postural activities was found. Resulting DEs and DPs differed from the TB conditions, especially in lower extremity activities. The differences between the percent changes of DEs vs. Vt found for lower extremity activities were confirmed by both ventilatory and postural diaphragm recruitment in response to postural demands.

Induced apoptosis of chondrocytes by Porphyromonas gingivalis as a possible pathway for cartilage loss in rheumatoid arthritis.

The role of bacterial infections in the pathogenesis of rheumatoid arthritis (RA) has gained increasing interest. Patients with RA often exhibit periodontal disease, which is associated with pathogens like Porphyromonas gingivalis. The present study examines the direct effects of P. gingivalis on apoptosis of human chondrocytes (a feature of inflammatory joint diseases) as one can assume an interrelation of pathogenesis of RA and P. gingivalis infections. Primary chondrocytes were infected with P. gingivalis. Early apoptotic and dead cell analysis was performed using Annexin-V, 7AAD, and propidium iodide and examined by flow cytometry and fluorescence microscopy. Caspase activation and DNA fragmentation were determined by western blot analysis and TUNEL reaction. Flow cytometry and fluorescence microscopy demonstrated an increase of Annexin-V-positive early apoptotic chondrocytes after infection. Western blot showed upregulation of activated caspase-3 expression, and TUNEL reaction revealed considerable DNA fragmentation following infection. The data show that P. gingivalis promotes early and later stages of apoptosis of primary human chondrocytes, which might contribute to the joint damage seen in the pathogenesis of RA.

Effects of hypoxia and/or lack of glucose on cellular energy metabolism and cytokine production in stimulated human CD4+ T lymphocytes.

Oxidative phosphorylation and/or glycolysis provide energy, mainly in the form of ATP, which ensures proper functioning of immune cells such as CD4(+) T lymphocytes. However, the main substrates, namely oxygen and glucose, are known to remain for a relatively short time in the inflamed tissue and in other clinical situations where immune cells need to function properly. Therefore, we examined the effect of hypoxia and/or lack of glucose on cellular energy metabolism and on cytokine secretion in stimulated human CD4(+) T lymphocytes. Human CD4(+) T cells were MACS-isolated using peripheral blood obtained from healthy donors. Stimulated cells were incubated in medium with or without glucose for 6h in a sealed chamber which led to cumulative hypoxia. During this incubation period, (i) oxygen saturation was measured continuously using a Clark-type electrode, and (ii) samples were taken at different time points in order to quantify for each the viability of cells, intracellular reactive oxygen species (iROS), ATP levels, glycolytic enzyme activity, mRNA expression of hexokinase-1 and superoxide dismutase-1, and concentrations of several different cytokines. Stimulated CD4(+) T cells which were incubated under normoxic conditions served as controls. Under hypoxic conditions, lack of glucose exerted a biphasic effect on cellular oxygen consumption: initially higher but later lower respiration rates were measured when compared to conditions where glucose was available. Lack of glucose strongly increased the number of dead cells and the formation of iROS under normoxia but not under hypoxia. Under both normoxic and hypoxic conditions, intracellular ATP levels remained almost unchanged during the incubation period if glucose was present, but decreased significantly in the absence of glucose, despite the enhanced glycolytic enzyme activity. Measurements of stimulated cytokine production demonstrated (i) that cumulative hypoxia stimulates especially the secretion of IL-1beta, IL-10 and IL-8, and (ii) that lack of glucose results in lower cytokine concentrations. We demonstrate that CD4(+) T cells are highly adaptive in bioenergetic terms which ensure their proper function under extreme conditions of glucose and/or oxygen availability as found under physiological and pathophysiological conditions. Hypoxia seems to facilitate inflammatory reactions and angiogenesis.

Intravitreal pegaptanib sodium for choroidal neovascularisation secondary to age-related macular degeneration: Pan-European experience.

PURPOSE: To evaluate visual outcomes in patients with neovascular age-related macular degeneration (NV-AMD) who were treated with pegaptanib sodium in European clinical ophthalmology practices. METHODS: Thirteen centres in eight European countries participated in this retrospective study. Medical records for patients with any angiographic subtype of subfoveal choroidal neovascularisation secondary to NV-AMD with visual acuities (study eye) of 20/40-20/320 treated with 0.3 mg pegaptanib as first-line treatment and with at least 24 weeks of follow-up were identified. Anonymised data reflecting at least 24 and up to 54 weeks of follow-up were recorded. Primary end points were visual acuity outcomes at weeks 24 and 54 compared with those reported at week 54 in the vascular endothelial growth factor (VEGF) Inhibition Study in Ocular Neovascularisation (VISION) trial. RESULTS: In all, 253 patients were followed for at least 24 weeks; 62 patients completed 54 weeks of follow-up. A mean of 4.4 (SD, 1.8) pegaptanib injections were administered through 24 weeks. Compared with the VISION trial, the European experience showed that >90% of patients in the current cohort lost <15 letters from baseline at both time points compared with 70% in the VISION trial at 54 weeks. Pegaptanib was well tolerated with no reported cases of endophthalmitis, traumatic cataract, or iatrogenic retinal detachment. CONCLUSIONS: Pegaptanib was found to stabilise vision in a greater percentage of patients and produced greater overall visual improvement in this group of treatment-naive patients with NV-AMD compared with outcomes reported in the VISION trial; however, interpretation of these results should be tempered given the differences in design between this retrospective study and the prospective controlled trial.

Study of cytotoxic effect of photodynamically and sonodynamically activated sensitizers in vitro.

High resolution imaging of biological structures and their changes induced by different agents such as drugs are commonly performed by confocal and electron microscopy. The past decade has witnessed an emersion of the atomic force microscopy (AFM) from solid-state physics into cell biology and even medical applications. For these reasons, we used this relatively new microscopic technique to study the morphology of cell lines. We imaged the cells by atomic force microscopy before and after the photodynamic therapy (PDT) using the photosensitizer ClAlPcS(2). We also compared the impact of the photosensitizer in combination with silymarin antioxidant on cancer and non-cancer cell lines by measuring the kinetic production of reactive oxygen species (ROS). PDT was induced by LED source with total irradiation dose of 15 J cm(-2) and SDT was induced by therapeutic ultrasound with frequency of 1 MHz, intensity 2 W cm(-2) and time of exposition 10 min. The results show ROS kinetic production within the cells during PDT, sonodynamic therapy (SDT) and modification of morphological features investigated by AFM. The combination of a sensitizer and the specific light source can lead to the loss of surface rigidity and eventually to dramatic changes of the cell shape, which we can study by AFM.

Effects of celecoxib on the expression of osteoprotegerin, energy metabolism and cell viability in cultured human osteoblastic cells.

BACKGROUND AND OBJECTIVE: The selective COX-2 inhibitor celecoxib is widely used to treat pain and inflammation in rheumatoid arthritis, osteoarthritis, and ankylosing spondylitis. The drug has well-known important effects on immune cells but its direct and/or indirect influence on osteoblasts has not yet been explored in detail. This study aimed to investigate the dose-dependent effects of celecoxib on cell viability, energy metabolism and bone remodeling processes in cultured human osteoblastic cells. METHODS: Primary human osteoblasts and MG-63 cells were incubated with celecoxib (2, 10, 50microM). Cell viability and apoptosis were determined by trypan blue, 7AAD and Annexin-V staining. Effects on cellular oxygen consumption were measured amperometrically using a Clark electrode. mRNA expression of GLUT-1 and OPG was determined by RT-PCR; OPG protein secretion by ELISA and HIF-1alpha protein expression by immunoblotting. RESULTS: While celecoxib at a concentration of 2 and 10microM showed only marginal effects, a suprapharmacological concentration of 50microM influenced viability and energy metabolism, as well as OPG expression and secretion of osteoblastic cells. Cell viability was significantly reduced by celecoxib treatment. Celecoxib at 50microM stimulated oxygen consumption significantly. Corresponding experiments with the protonophore FCCP suggest that this effect is due to mitochondrial uncoupling. After 24h, GLUT-1 mRNA expression was significantly increased. HIF-1alpha protein was not expressed under any of our experimental conditions. We also showed that celecoxib at 50microM significantly inhibits OPG protein secretion leading to a compensative increase of mRNA expression. CONCLUSION: Pronounced effects of celecoxib on cell viability (reduction), oxygen consumption (stimulation), GLUT-1 mRNA expression (stimulation) and OPG protein secretion (inhibition) in osteoblastic cells were observed only at 50microM-a concentration not reached by therapeutic doses giving plasma concentrations less than 10microM. On the contrary, celecoxib at 2 and 10microM showed only marginal effects, suggesting that celecoxib administration is probably safe with respect to bone metabolism in cases requiring potent treatment of pain and inflammation. However, higher intracellular concentrations, which might occur through accumulation, necessitate investigations with high concentrations.

Analysis of diaphragm movement during tidal breathing and during its activation while breath holding using MRI synchronized with spirometry.

Using magnetic resonance imaging (MRI) in conjunction with synchronized spirometry we analyzed and compared diaphragm movement during tidal breathing and voluntary movement of the diaphragm while breath holding. Breathing cycles of 16 healthy subjects were examined using a dynamic sequence (77 slices in sagittal plane during 20 s, 1NSA, 240x256, TR4.48, TE2.24, FA90, TSE1, FOV 328). The amplitude of movement of the apex and dorsal costophrenic angle of the diaphragm were measured for two test conditions: tidal breathing and voluntary breath holding. The maximal inferior and superior positions of the diaphragm were subtracted from the corresponding positions during voluntary movements while breath holding. The average amplitude of inferio-superior movement of the diaphragm apex during tidal breathing was 27.3+/-10.2 mm (mean +/- SD), and during voluntary movement while breath holding was 32.5+/-16.2 mm. Movement of the costophrenic angle was 39+/-17.6 mm during tidal breathing and 45.5+/-21.2 mm during voluntary movement while breath holding. The inferior position of the diaphragm was lower in 11 of 16 subjects (68.75 %) and identical in 2 of 16 (12.5 %) subjects during voluntary movement compared to the breath holding. Pearson's correlation coefficient was used to demonstrate that movement of the costophrenic angle and apex of the diaphragm had a linear relationship in both examined situations (r=0.876). A correlation was found between the amplitude of diaphragm movement during tidal breathing and lung volume (r=0.876). The amplitude of movement of the diaphragm with or without breathing showed no correlation to each other (r=0.074). The movement during tidal breathing shows a correlation with the changes in lung volumes. Dynamic MRI demonstrated that individuals are capable of moving their diaphragm voluntarily, but the amplitude of movement differs from person to person. In this study, the movements of the diaphragm apex and the costophrenic angle were synchronous during voluntary movement of the diaphragm while breath holding. Although the sample is small, this study confirms that the function of the diaphragm is not only respiratory but also postural and can be voluntarily controlled.

Fat embolism syndrome.

Fat embolism syndrome is a rare complication occurring in 0.5 to 2% of patients following a long bone fracture. It is believed to be caused by the toxic effects of free fatty acids. Diagnosis is clinical, based on respiratory, cerebral and dermal manifestations. Treatment is only supportive, directed mainly at maintaining respiratory functions.