Evaluation of blood cell attachment on Er: YAG laser applied root surface using scanning electron microscopy.

BACKGROUND: Periodontal regeneration is dependent on the uninterrupted adhesion, maturation and absorption of fibrin clots to a periodontally compromised root surface. The modification of the root surface with different agents has been used for better fibrin clot formation and blood cell attachment. It is known that Er:YAG laser application on dentin removes the smear layer succesfully. AIM: The aim of this study is to observe blood cell attachment and fibrin network formation following ER:YAG laser irradiation on periodontally compromised root surfaces in comparison to chemical root conditioning techniques in vitro. MATERIALS AND METHODS: 40 dentin blocks prepared from freshly extracted periodontally compromised hopeless teeth. Specimens were divided in 5 groups; those applied with PBS, EDTA, Citric acid and Er:YAG. They were further divided into two groups: those which had received these applications, and the control group. The specimens were evaluated with scanning electron microscope and micrographs were taken. Smear layer and blood cell attachment scoring was performed. RESULTS: In the Er:YAG laser applied group, smear layer were totally removed. In the blood applied specimens, better fibrin clot formation and blood cell attachment were observed in the Er:YAG group. In the group that had been applied with citric acid, the smear layer was also removed. The smear layer could not be fully removed in the EDTA group. CONCLUSION: Er:YAG laser application on the root dentin seems to form a suitable surface for fibrin clot formation and blood cell attachment. Further clinical studies to support these results are necessitated.

Glutamic acid decarboxylase immunoreactivity in the mossy fiber terminals of the hippocampus of genetic absence epileptic rats.

AIM: Genetic absence epilepsy rats from Strasbourg (GAERS) provide a model of absence epilepsy. Although excessive GABA mediation within the thalamo-cortico-thalamic circuit has been shown to play a role in absence epilepsy, neuronal networks of hippocampus have recently received attention. Glutamic acid decarboxylase (GAD) was previously shown to be increased after convulsive seizures in the mossy fiber terminals (MFTs) of hippocampus. The aim of the present study was to investigate whether the change in the level of this enzyme in convulsive seizures is also observed in rats having genetic absence epilepsy. MATERIAL AND METHODS: Hippocampal CA3 and dentate regions were processed for transmission electron microscopic evaluations. Thin sections were incubated with anti-GAD65/67 antibody. The NIH Image Analysis program was used for the quantitative analysis. RESULTS: It was observed that GAD65/67 immunoreactivity was positive in CA3 and dentate gyrus MFTs of both groups and the difference in the density of immunolabeling between the groups was not statistically significant. CONCLUSION: The present study demonstrated that GABA synthesizing enzyme, GAD, is found in MFTs of Wistar and GAERS hippocampus and this enzyme does not show an increase in these terminals in absence epilepsy, in contrast to convulsive seizures.

Effect of extremely low frequency electromagnetic fields on growth rate and morphology of bacteria.

PURPOSE: To determine the effect of extremely low frequency (<300 Hz) electromagnetic fields (ELF-EMF) on the growth rate of Gram-positive and Gram-negative bacteria and to determine any morphological changes that might have been caused by ELF-EMF. MATERIALS AND METHODS: Six bacterial strains, three Gram-negative and three Gram-positive were subjected to 50 Hz, 0.5 mT ELF-EMF for 6 h. To determine growth rate after ELF-EMF application, bacteria exposed to ELF-EMF for 3 h were collected, transferred to fresh medium and cultured without field application for another 4 h. Growth-rate was determined by optical density (OD) measurements made every hour. Morphological changes were determined with Transmission electron microscopy (TEM) for two gram-negative and two gram-positive strains collected after 3 h of field application. RESULTS: A decrease in growth rate with respect to control samples was observed for all strains during ELF-EMF application. The decrease in growth-rate continued when exposed bacteria were cultured without field application. Significant ultrastructural changes were observed in all bacterial strains, which were seen to resemble the alterations caused by cationic peptides. CONCLUSION: This study shows that ELF-EMF induces a decrease in growth rate and morphological changes for both Gram-negative and Gram-positive bacteria.

Ultrastructural GABA immunocytochemistry in the mossy fiber terminals of Wistar and genetic absence epileptic rats receiving amygdaloid kindling stimulations.

The existence of absence epilepsy and temporal lobe epilepsy in the same patient is not common in clinical practice. The reason why both types of seizures are rarely seen in the same patient is not well understood. Therefore, we aimed to investigate kindling in a well known model of human absence epilepsy, genetic absence epilepsy rats from Strasbourg (GAERS). In the present study, we analyzed whether the GABA content of GAERS that received kindling stimulations was altered in the hippocampal mossy fiber terminals compared to non-epileptic control (NEC) Wistar rats. For this purpose, we used an immunocytochemical technique at the ultrastructural level. Ultrathin sections were immunolabeled with anti-GABA antibody and transmission electron microscopy was used for the ultrastructural examination. The number of gold particles per nerve terminal was counted and the area of the nerve terminal was determined using NIH image analysis program. The GABA density was found to be higher in sham-operated GAERS than sham-operated Wistar rats. The density was increased in kindling Wistar group compared to sham-operated Wistar and kindling GAERS groups. No statistical difference was observed between sham-operated GAERS and kindling GAERS groups. The increase in GABA levels in stimulated Wistar rats may be a result of a protective mechanism. Furthermore, there may be strain differences between Wistar rats and GAERS and our findings addressing different epileptogenesis mechanisms in these strains might be a basis for future experimental studies.

Quantitative analysis of ciliary ultrastructure in patients with primary ciliary dyskinesia.

The present study was designed to investigate dynein arm and microtubule defects quantitatively in patients with respiratory disease and to establish the clinical relevance of dynein arm deficiency and microtubule abnormalities. Thirty-four patients with recurrent upper and/or lower respiratory infections were included in the study. Nasal mucosal brushings were fixed in glutaraldehyde and routine electron microscopic procedures were carried out. At least 20 cross-sectioned cilia were examined from each subject. Dynein arm and microtubular abnormalities were quantified and a statistical analysis was performed. Twenty-nine percent of the patients showed dynein arm deficiency and a further 21% had possible deficiency (PD). Microtubule defects in patients with dynein arm deficiency and PD were found to be significantly increased compared to the patients with no dynein arm deficiency. The most prominent defect in the dynein arm deficiency group was a translocation of central and/or peripheral microtubules. The high percentage of translocation defect in this group of patients suggests that these defects are primary, rather than secondary to infection.

Anti-inflammatory effect of meloxicam on experimental vasospasm in the rat femoral artery.

Cerebral vasospasm influences morbidity and mortality following subarachnoid haemorrhage (SAH). Inflammation is believed to play a role in post-haemorrhagic vasospasm. Meloxicam is a non-steroidal anti-inflammatory drug. We investigated the effect of meloxicam on a rat femoral artery vasospasm model using the radial wall thickness and cross-sectional lumen area as parameters under light, scanning and transmission electron microscopy examination. Rats were randomly separated into SAH, SAH+ meloxicam and control groups. Rats in the SAH+ meloxicam group were given meloxicam at 2 mg/kg daily for 7 days. Femoral arteries were examined by light microscopy and scanning and transmission electron microscopy, and for morphometric analysis. A statistically significant difference (p<0.001) was detected between the SAH and SAH+ meloxicam groups. Meloxicam treatment reduced ultrastructural and morphometric vasospastic changes. These findings support the hypothesis that inflammation may play a role in the pathophysiologyical pathways of post-haemorrhagic cerebral vasospasm.

The effect of temporary aneurysm clip on the common carotid artery of atherosclerotic rabbits.

BACKGROUND: We compared the effect of temporary aneurysm clips on atherosclerotic and nonatherosclerotic CCA of rabbits by morphometric and ultrastructural methods. METHODS: The rabbits (N = 12) were divided into 2 groups: the first group was fed a 2% cholesterol diet, and the second group, a normal diet for 4 weeks. Atherosclerotic lesions developed after 4 weeks. Temporary aneurysm clips were placed on the left CCA of both groups; the right CCA of both groups served as control. Thus, a total of 4 groups were used: atherosclerotic (A), atherosclerotic/clip (AC), nonatherosclerotic (NA), and nonatherosclerotic/clip (NAC). Temporary aneurysm clips were applied for 1, 5, and 10 minutes in the AC and NAC groups. No temporary clip was placed on the right CCA (A and NA groups). The affected parts of the CCA via clips were examined under light microscope and SEM. RESULTS: Comparison of atherosclerotic and nonatherosclerotic CCA of rabbits under light microscope indicated that the wall of atherosclerotic CCA was thicker than that of nonatherosclerotic CCA. The difference between the thickness of atherosclerotic and nonatherosclerotic CCAs was significant. SEM analyses showed that in nonatherosclerotic CCAs, the effect of temporary aneurysm clips was seen after 10 minutes, but in atherosclerotic CCAs, the effect was seen within the 1st minute of clipping and continued in the 5th and 10th minutes. CONCLUSION: The duration of temporary clipping should be decreased for the neurovascular surgery of atherosclerotic patients.

Time-dependent changes in distribution of basic fibroblast growth factor immunoreactive cells in rat hippocampus after status epilepticus.

OBJECTIVE: In this study, we aimed to examine time-dependent morphologic changes and quantitative alterations in the density of basic fibroblast growth factor (bFGF)-immunoreactive (ir) astrocytes and CA2 pyramidal neurons in dorsal hippocampus of rats after status epilepticus (SE) induced by kainic acid (KA) injection. METHODS: Wistar albino rats were injected with saline or KA i.p. to investigate time-dependent alterations in morphology and the number of bFGF-ir astrocytes and neurons in the dorsal hippocampus 15, 30 and 90 days after KA injection. RESULTS: Fifteen days after KA injection, gliosis was present throughout the hippocampus and neuronal loss was evident in CA1 and CA3 regions, which was more severe after 30 and 90 days. KA-injected rats demonstrated significantly increased number of both bFGF-ir astrocytes throughout the hippocampus and pyramidal neurons in CA2 after 15 days and decreased number after 30 and 90 days. CONCLUSION: The decrease in the number of bFGF-ir astroglia and neurons in long term after KA injection may indicate a decrease in the production of bFGF and/or number of bFGF-ir cells, suggesting that protective effects of bFGF might be altered during epileptogenesis in the hippocampus.

Time-dependent changes in distribution of basic fibroblast growth factor immunoreactive cells in hippocampus after kainic acid injection in rat pups.

Five-day-old Wistar albino rats were injected with kainic acid (KA) or saline i.p. to investigate time-dependent alterations in morphology and number of basic fibroblast growth factor (bFGF) immunoreactive (-ir) astrocytes and neurons in hippocampus at 15, 30, and 90 days after the injections. Sections were stained with cresyl violet for morphological evaluation and bFGF immunohistochemistry was used for quantitative evaluation of bFGF-ir cell density. Fifteen days after KA injection, there was gliosis but no neuronal loss although disorganization in CA1, CA3, CA4 pyramidal layers and neuronal loss were evident 30 and 90 days after the injection. KA injected rats demonstrated significantly increased number of bFGF-ir astrocytes throughout the hippocampus and pyramidal neurons in CA2 after 15 days and decreased number of bFGF-ir cells after 30 and 90 days. The decrease in the number of bFGF-ir astroglia and neurons in long term after KA injection may indicate a decrease in the production of bFGF and/or number of bFGF-ir cells suggesting that protective effects of bFGF may be altered during epileptogenesis in hippocampus.

Evaluation of hexagonal boron nitride as a new tablet lubricant.

In this study, hexagonal boron nitride (HBN) was evaluated as a new lubricant for pharmaceutical tablet manufacturing. The other conventional lubricants such as magnesium stearate (MGST), stearic acid (STAC), and glyceryl behenate (COMP) were also tested along with HBN. Tablets were manufactured on an instrumented single-station tablet press to monitor and quantify the lower punch ejection force (LPEF). The force ratio, tablet crushing strength, disintegration time, and thickness were measured. The lubricant film formation and lubricant distribution in tablets were studied using the scanning electron microscopy (SEM) and electron probe micro analyzer (EPMA). Based on the force ratio, a good lubrication was obtained at 1% for MGST and HBN; in contrast, STAC and COMP did not show a good lubrication. After 1%, all lubricants performed well. MGST was found to be the most effective lubricant based on LPEF-lubricant concentration profile. HBN provided a 50% decrease in LPEF at 2% lubricant concentration and was rated as an effective tablet lubricant. HBN was better than either STAC or COMP. Unlike MGST, HBN had no significant prolongation effect on tablet disintegration times.

Glutamate and GABA immunocytochemical electron microscopy in the hippocampal dentate gyrus of normal and genetic absence epilepsy rats.

It is generally accepted that absence epilepsy results from the impairment of GABAergic and glutamatergic neurotransmission. In particular, besides excessive GABA mediation within the thalamo-cortico-thalamic circuit in absence epilepsy, neuronal networks of the hippocampus have recently received attention. In the present study, we examined the density of glutamate and GABA neurotransmitter immunolabeling in the dentate gyrus of the hippocampus of genetic absence epilepsy rats from Strasbourg (GAERS) compared to the control group. GABA and glutamate were found to exist in synaptic vesicles of the mossy fiber terminals of the control and GAERS groups. The density of glutamate immunolabeling within the mossy fiber terminals in the hilar region of GAERS hippocampus was found to be significantly decreased compared to the control group. There was no difference in the density of immunolabeling within GABA nerve terminals between GAERS and control group. The findings of this study suggest that mechanisms underlying absence seizures in GAERS may also manifest themselves in other brain regions such as the hippocampus. The presence of GABA within synaptic vesicles of mossy fiber terminals, as revealed by high resolution ultrastructural immunocytochemistry, has provided additional evidence to the possible modulatory role of GABA on synaptic transmission between the mossy fiber and the target cell.

Collagen ultrastructure and TGF-beta1 expression preserved with aminoguanidine during wound healing in diabetic rats.

Advanced glycoxidation end products have been implicated in delayed diabetic wound healing. In this study, we evaluated the effects of aminoguanidine, which is an advanced glycation and nitric oxide (NO) synthase inhibitor, on extracellular matrix protein expression, collagen configuration, and nitrite/nitrate levels in wounds of diabetic rats. Sixteen Wistar male rats were made diabetic by streptozotocin. Of these, eight rats were given AG (aminoguanidine bicarbonate (AG) (group DAG) in their drinking water, and eight rats were followed as diabetic paired controls (group D). Eight healthy rats were followed as the healthy control group (group H). At the eighth week, a 2 x 2 cm area full-thickness skin defect was created. The degree of contraction of the open wounds was evaluated for 2 weeks duration. On the 15th postoperative day, wound surface areas were measured, and wound specimens and blood samples were collected. The shrinking percentage of the wounds was small in both groups H and DAG compared with group D (p < 0.05). Similar to healthy rats, the aminoguanidine-treated diabetic rats had very strong transforming growth factor (TGF)-beta1 expression in granulation tissue and intact skin in comparison with diabetic controls. In the diabetic group, the intact skin demonstrated sparsely distributed regular collagen fibers in the granulation zone, and the regular pattern of collagen fibers was lost. In conclusion, aminoguanidine improves wound healing, restores growth factor TGF-beta1 expression, and preserves collagen ultra structure, whereas it has no prominent effect on NO levels within wound tissue in diabetic rats.

Connections between the facial, vestibular and cochlear nerve bundles within the internal auditory canal.

The vestibular, cochlear and facial nerves have a common course in the internal auditory canal (IAC). In this study we investigated the average number of nerve fibres, the average cross-sectional areas of the nerves and nerve fibres, and the apparent connections between the facial, cochlear and vestibular nerve bundles within the IAC, using light and scanning electron microscopy. The anatomical localization of the nerves within the IAC was not straightforward. The general course showed that the nerves rotated anticlockwise in the right ear from the inner ear end towards the brainstem end and vice versa for the left ear. The average number of fibres forming vestibular, cochlear, and facial nerves was not constant during their courses within the IAC. The superior and the inferior vestibular nerves showed an increase in the number of nerve fibres from the inner ear end towards the brainstem end of the IAC, whereas the facial and the cochlear nerves showed a reduction in the number of fibres. This suggests that some of the superior and inferior vestibular nerve bundles may receive fibres from the facial and/or cochlear nerves. Scanning electron microscopic evaluations showed superior vestibular-facial and inferior vestibular-cochlear connections within the IAC, but no facial-cochlear connections were observed. Connections between the nerves of the IAC can explain the unexpected vestibular disturbances in facial paralysis or persistence of tinnitus after cochlear neurectomy in intractable tinnitus cases. The present study offers morphometric and scanning electron microscopic data on the fibre connections of the nerves of the IAC.

Simple and reliable detection of slime production of Candida spp. directly from blood culture bottles: comparison of visual tube method and transmission electron microscopy.

Early detection of slime production may be useful for clinical decision because of its suggestive property for potential pathogenic capacity of a Candida strain especially in patients with a prosthetic device. In this study we aimed to compare the visual tube method (VTM) with transmission electron microscopy (TEM) in order to confirm the reliability of the former method. In order to demonstrate the reproducibility of the tube method and to determine the correct timing for the test, Candida isolates directly obtained from blood culture (DBC) bottles and their two subsequent subcultures were used. The results of this study showed that VTM is a simple and reliable method which can be used in every clinical mycology laboratory, provided that the test is applied on DBC isolates; as the ability of slime production is decreased or lost even after the first subculturing. We suggest that this simple method can be used and may have some contributions to the ongoing studies on the controversial issue concerning removal of biomaterials in candidemic patients.

Immunocytochemical analysis of glutamate and GABA in hippocampus of genetic absence epilepsy rats (GAERS).

In the present study, we used an immunocytochemical technique at the electron microscopic level to determine if there are changes in the glutamate and GABA neurotransmitter content of the hippocampus of genetic absence epilepsy rats from Strasbourg (GAERS). We also investigated if there was mossy fiber reorganization. After perfusion fixation, brains were removed and cryostat sections were stained according to the neo-Timm's procedure. High-resolution electron microscopy was used for ultrastructural examination of the hippocampus of GAERS and non-epileptic control Wistar animals. For ultrastructural and immunocytochemical studies, ultrathin-cut sections were obtained and immunolabeled with anti-glutamate and anti-GABA antibodies. The number of gold particles per nerve terminal was counted and the area of the nerve terminal was determined using the program NIH Image Analysis. No mossy fiber sprouting was detected in the hippocampus of GAERS. GABA and glutamate immunoreactivity were observed in the mossy fiber terminals of both the control and GAERS groups. Glutamate density in the CA3 region of GAERS hippocampus was found to be significantly increased compared to the control group. However, there was no difference in the GABA density of nerve terminals and in areas of GABAergic and mossy terminals between GAERS and the control group. The difference in glutamate level may merely be due to strain differences between the GAERS strain and the original Wistar strain or it is also possible that it appears after seizures have started.

The ameliorating effect of melatonin on protamine sulfate induced bladder injury and its relationship to interstitial cystitis.

PURPOSE: The pineal hormone melatonin was recently shown to have free radical scavenging ability and it reduces lipid peroxidation. In this morphological study we investigated the effects of melatonin on protamine sulfate (Sigma Chemical Co., St. Louis, Missouri) induced bladder injury. MATERIALS AND METHODS: Albino Wistar female rats were catheterized and intravesically infused with phosphate buffered solution (control group) or protamine sulfate (bladder injury group) dissolved in phosphate buffered solution. In the protamine sulfate plus melatonin group after protamine sulfate instillation melatonin was injected intraperitoneally. Bladder morphology was investigated by light and electron microscopy. Tissue samples were also obtained to determine bladder malondialdehyde levels. RESULTS: In the bladder injury group ulcerated areas, an irregular glycosaminoglycan layer, increased number of mast cells, vacuole formation, dilated perinuclear cistern, formation of pleomorphic and uniform microvilli, and dilated urothelial intercellular spaces were observed. In the bladder injury plus melatonin group a relatively normal urothelial topography, glycosaminoglycan layer and decreased number of mucosal mast cells, some dilatation between intercellular areas, less uniform microvilli and in most areas regular tight junctions were observed. CONCLUSIONS: Increased malondialdehyde levels as a result of protamine sulfate induction lead us to propose that free radicals may have a critical role in this injury. The significant decrease in malondialdehyde levels in the protamine sulfate plus melatonin group was in accordance with morphological findings. Thus, melatonin appears to exert a urothelial protective activity in a bladder injury model.

Connections of the dorsomedial hypothalamic nucleus from the forebrain structures in the rat.

The dorsomedial hypothalamic nucleus (DMH) has been implicated as an area controlling autonomic activity. The aim of this study was to demonstrate connections of the anterior and posterior DMH to the forebrain structures, using a horseradish peroxidase (HRP) retrograde axonal transport technique in rats. The results of HRP labelling show that the anterior and posterior DMH indicate a number of differences in their connections. The posterior DMH has intense connections with the cortex (cingulate, frontal, parietal and insular), amygdala (lateral and basolateral) and hippocampus (CA1 and CA2), whereas the anterior DMH has faint connections with the cortex (cingulate, frontal and parietal) and prominent connections with the septal and bed nucleus of stria terminalis. These differences in connections of the DMH may provide sites for the specific autonomic function integrated by the DMH.