Evaluation of cell morphology and adhesion capacity of human gingival fibroblasts on titanium discs with different roughened surfaces: an in vitro scanning electron microscope analysis and cell culture study.

BACKGROUND: Implantoplasty is an option in peri-implantitis treatment. What is known about the effects of implantoplasty on peri-implant soft tissue adhesion and cell behaviours is limited. This study aimed to evaluate the morphological features and adhesion capacity of human gingival fibroblast (HGF) cells onto sand-blasted, large-grit, acid-etched (SLA®) titanium (Ti) discs surfaces roughened with different implantoplasty protocols. MATERIALS AND METHODS: The study included a total of 48 Ti discs divided into four groups (n = 12 per group): Group I: machined, smooth surface discs; Group II: SLA® surface discs; Group III: SLA® surface discs roughened with diamond bur sequence (40 and 15-µm grit); Group IV: SLA® surface discs roughened with diamond bur sequence (125 and 40-µm grit). Following polishing procedure, the surface roughness value of discs was assessed by a profilometer and scanning electron microscope. HGFs were cultured on Ti discs and cell adhesion was examined after the 24th, 48th, and 72nd hours. Statistical significance was set at the p ≤ 0.05 level. RESULTS: Scanning electron microscope analyses of the discs revealed that fibroblasts exhibited well-dispersion and a firm attachment in all groups. The cells in group I and II had thin and long radial extensions from the areas where the nucleus was located to the periphery; however, attached cells in group III and IV showed more spindle-shaped morphology. The surface roughness parameters of the test groups were lower than those of the SLA®. The SLA® group showed the highest HGF adhesion (group II) (p ≤ 0.05). HGF adhesion in group IV was greater compared to group III, but less than group I. CONCLUSIONS: This study showed that the characteristics of the burs applied in the implantoplasty protocol are determinant for the surface roughness and fibroblast adhesion occurs on surfaces with decreased roughness following implantoplasty. Consequently, it should be kept in mind that the surface properties of the implant may affect the adherent cell morphology and adhesion.

The cardioprotective effects of thoracal epidural anestesia are induced by the expression of vascular endothelial growth factor and inducible nitric oxide synthase in cardiopulmonary bypass surgery.

AIM: The cardioprotective effects of thoracal epidural anesthesia (TEA) are induced by the expression of vascular endothelial growth factor (VEGF) and inducible nitric oxide synthase (i-NOS) in cardiopulmonary bypass (CPB) surgery. When general anaesthesia (GA) is combined with TEA during coronary artery bypass graft, we investigated whether TEA together with GA play a role on VEGF and i-NOS expression in human heart tissue in cardiac ischemia. METHODS: Right atrial biopsy samples were taken before CPB, before aortic cross clamp (ACC) and at 15 min after ACC release (after ischemia and reperfusion). Human heart tissues were obtained from the TEA+GA and GA groups. Immunocytochemistry was performed using antibodies for VEGF and i-NOS. RESULTS: Both VEGF and i-NOS immunoreactivity was observed in cardiomyocytes and arteriol walls. Although VEGF and i-NOS immunoreactivity was apparent in both groups,, immunostaining intensity was greater in the TEA+GA group than the GA group. Between groups, at 4 h and at 24 h after the end of CPB, the cardiac index (CI) was significantly higher in the TEA+GA group than GA group (3.4+/-0.8 L/min/m(2) vs 2.5+/-0.8 L/min/m(2); P<0.001), (3.8+/-1.1 L/min/m(2) vs 3.1+/-1.1 L/min/m(2); P<0.008) respectively. Within groups, at 4 and 24 h after the end of CPB, the CI was significantly higher in the TEA+GA group than baseline values, (3.4+/-0.8 L/min/m(2) vs 2.4+/-0.7 L/min/m(2); P<0.001), (3.8 +/-1.1 L/min/m(2) vs 2.4+/-0.7 L/min/m(2); P<0.001) respectively, but no difference was found in the GA group (2.6+/-0.8 L/min/m(2) vs 2.5+/-0.8 L/min/m(2); P>0.05), (2.6+/-0.8 L/min/m(2) vs 3.1+/-1.1 L/min/m(2); P>0.05) respectively. After ACC release, 11/40 (27.5%) patients in the TEA+GA group showed ventricular fibrillation (VF), atrial fibrillation or heart block versus 25/40 (62.5%) of those in the GA group. VF after ACC release in the TEA+GA group (9/20 patients, 22.5%) was significantly lower than in the GA group (21/40 patients, 52.5%); (P<0.006). Sinus rhythm after ACC release in the TEA+GA group (29/40 patients, 72.5%) was significantly higher than in the GA group (15/40 patients, 37.5%); (P<0.002). CONCLUSIONS: The results of the present study indicate that TEA plus GA in coronary surgery preserve cardiac function via increased expression of VEGF and i-NOS, improved hemodynamic function and reduced arrhythmias after ACC release.

General anesthesia with thoracic epidural anesthesia in the cardiopulmonary bypass surgery reduces apoptosis by upregulating antiapoptotic protein Bcl-2.

AIM: The aim of the paper was to investigate whether thoracic epidural anesthesia (TEA) together with general anaesthesia (GA) play a role on apoptosis in humans before cardiopulmonary bypass (CPB), before aortic cross clamp (ACC) and at 15 min after ACC release (after ischemia and reperfusion). METHODS: Eighty patients scheduled for elective CABG were randomized to receive either GA group (n: 40) or TEA+GA group (n: 40). The right atrial biopsy samples were taken before CPB, before ACC and at 15 min after ACC release from all patients. Human heart tissues were obtained from patients of TEA+GA group and GA group. The number of Bcl-2 positive cardiomyocytes was counted in multiple tissue sections of biopsies of 80 patients using light microscopy (magnification x 40) with an ocular micrometer system (Olympus). RESULTS: In the TEA+GA group, the Bcl-2 positive cardiomyocytes were distinctly statistically increased compared to the GA group (P<0.001). In addition, the intensity of the immunostaining was also increased in the TEA+GA compared with the GA group. The number of immunoreactive cardiomyocytes is as follows: before CPB, TEA+GA group 396+/-61, GA group 92+/-41, before ACC, TEA+GA group 333+/-47, GA group 94+/-18, at 15 min after ACC release, TEA+GA group 346+/-68.8, GA group 85+/-9.5. There were statistically significant differences between groups, (P<0.001). Between groups, at 4 h and at 24 h after the end of CPB, in the TEA+GA group, the CI was significantly higher than GA group respectively; (3.4+/-0.8 L/min/m(2) vs 2.5+/-0.8 L/min/m(2); P<0.001), (3.8+/-1.1 L/min/m(2) vs 3.1+/-1.1 L/min/m(2); P<0.008). Within groups, at 4 and 24 h after the end of CPB, in the TEA+GA group, the CI was significantly higher than baseline values, respectively, (3.4+/-0.8 L/min/m(2) vs 2.4+/-0.7 L/min/m(2); P<0.001), (3.8+/-1.1 L/min/m(2) vs 2.4+/-0.7 L/min/m(2); P<0.001). Whereas no difference was found in the GA group respectively, (2.6+/-0.8 L/min/m(2) vs. 2.5+/-0.8 L/min/m(2); P>0.05), (2.6+/-0.8 L/min/m(2) vs. 3.1+/-1.1 L/min/m(2); P>0.05). The number of patients showing ventricular fibrillation (VF), atrial fibrillation or heart block after release of the ACC was 11 of 40 (27.5%) in the TEA+GA group versus 25 of 40 (62.5%) in the GA group. The number of patients showing VF after release of ACC was 9 out of 20 patients (22.5%) in the TEA+GA group which was significantly lower than in the GA group (21 of 40 patients 52.5%); (P<0.006). Sinus rhythm after release of ACC, in the TEA+GA group was observed in 29 of 40 patients (72.5%) and was significantly higher than in the GA group (15 of 40 patients 37.5%); (P<0.002). CONCLUSION: The result of the present study indicate that TEA plus GA in coronary surgery had preserved cardiac function during intraoperative and postoperative period by means of reduced apoptosis, improved hemodynamic function and reduced arrhythmias after release of the ACC.

Growth hormone-releasing hormone peptide and mRNA are overexpressed in GH-deficient Ames dwarf mice.

Hypothalamic expression of growth hormone-releasing hormone (GHRH) was quantified morphologically in dwarf mice which exhibit spontaneous genetic GH absence. Mouse GHRH mRNA was assessed by in situ hybridization; densitometric evaluation of total mRNA in dwarfs showed levels 2.3-fold higher than in phenotypically normal siblings (p < 0.01); assessment of mRNA per neuron by autoradiographic grain counting indicated a 2.5-fold increase per cell in dwarfs (p < 0.005). GHRH peptide was evaluated immunocytochemically using a new mouse-specific antiserum; numbers of neurons containing detectable levels were 3-fold higher in dwarfs (p < 0.005). The increase in GHRH mRNA corroborates that reported previously in the GH-deficient little mouse, and after hypophysectomy in rats; GHRH peptide increase contrasts with previous reports of the effect of acute GH removal by hypophysectomy, in which GHRH levels fell. The results suggest that chronic GH deficiency is accompanied by increased translation as well as transcription of GHRH.

Median eminence-afferent vasoactive intestinal peptide (VIP) neurons in the hypothalamus: localization by simultaneous tract tracing and immunocytochemistry.

Retrograde tract tracing and immunocytochemistry were used to investigate the CNS source of the VIP that is present in high concentrations in the hypophysial portal blood and has been shown to have a stimulatory effect on pituitary prolactin secretion. Fluoro-gold (FG), which enters the CNS through areas devoid of the blood-brain barrier, such as median eminence, was injected peripherally. Brain sections from FG-treated animals were immunostained for VIP. A small population of VIP-containing cell bodies in the parvocellular and periventricular parts of the paraventricular nucleus (PVN) was also labeled with FG. Vasoactive intestinal peptide-immunoreactive perikarya not labeled with FG were also observed in the PVN, as well as FG-labeled cells that did not contain VIP. The results suggest that some VIP-producing neurons in the PVN project to the median eminence and are, therefore, functionally related to pituitary regulation; the function of other VIP neurons in the PVN is unknown.

The cellular localization of vasoactive intestinal peptide (VIP) in the mouse median eminence by immuno-electron microscopy.

The aim of this study was to examine, by use of pre-embedding immunocytochemistry, the ultrastructural localization of vasoactive intestinal peptide (VIP) immunoreactivity in the mouse median eminence. VIP immunoreactivity was observed in axonal profiles. The VIP-immunoreactive axonal profiles were in close proximity to non-immunoreactive axonal profiles that contained dense granular vesicles and clear vesicles and also to processes of tanycytes. VIP-immunoreactive terminals were observed in the proximity of the perivascular space and in the neuropil. Our results suggest that VIP-immunoreactive axon terminals may possibly interact with other non-immunoreactive axon terminals containing peptide and/or other transmitters at the level of the median eminence or may be released to the portal vasculature thereby to effect anterior pituitary cells.

Antiproliferative effects and insulin-like growth factor-I expression in Balb-C 3T3 fibroblasts after treatment with somatostatin and gonadotropin-releasing hormone analogs.

The present study was aimed to compare antiproliferative effects of somatostatin (SS) and gonadotropin-releasing hormone analogs (GnRHa) on a fibroblast cell line. Proliferation index, cell count, viability of the cells and insulin-like growth factor-I (IGF-I) immunoreactivity were determined after treatment with either SS (100 microM/ml), GnRHa (35 nM/ml) or SS and GnRHa of Balb-C 3T3 mouse fibroblasts. It was found that the proliferation index, cell count, viability and IGF-I immunoreactivity were not affected by GnRHa treatment as compared with no treatment (p > 0.05). Application of SS to the fibroblasts resulted in a significant reduction in proliferation index, cell count, and IGF-I immunoreactivity as compared with GnRHa treatment and no treatment, but it had no effect on cell viability. The labelling index in SS-treated cells was significantly reduced as compared with combined treatment with SS and GnRHa. In conclusion, a direct effect of GnRHa on fibroblast cells in culture could not be demonstrated. SS had direct inhibitory effects on cell proliferation possibly via inhibition of IGF-I effects without affecting cell viability.

Expression of insulin-like growth factor in the placenta of intrauterine growth-retarded human fetuses.

Many cases of intrauterine growth retardation (IUGR) are the result of placental and fetal tissue insufficiency. Insulin-like growth factor-I (IGF-I) is known to play a role in placental and fetal growth. An immunocytochemical study was performed to localize IGF-I peptides in human placenta and umbilical cords of normal (n = 3) and IUGR (n = 3) fetuses. The peripartum fetal conditions were evaluated as well. Immunoreactive IGF-I was detected in the cytotrophoblast, syncytiotrophoblast, amnion, endothelial cells of fetal capillaries and in the decidua in both normal and IUGR placental tissue. A more robust immunostaining and increased numbers of positively stained cells were found in the decidua of IUGR placenta (p < 0.001). Intense immunostaining was also found in endothelial cells, smooth muscle cells and fibroblasts of the umbilical vein. IGF-I immunoreactivity was also present in stroma (Hofbauer cells and/or fibroblasts) of IUGR villi. Our results indicate that expression of IGF-I is high in specific sites in placenta and umbilical cords, which indicates a paracrine and/or endocrine function. The increased expression of IGF-I in placenta of IUGR fetuses indicates its involvement in restoring normal growth by means of a positive feed-back mechanism.

Localization of neural cell adhesion molecule (N-CAM) immunoreactivity in adult rat tissues.

Neural cell adhesion molecule (N-CAM) mediates homophilic adhesion between cells and heterophilic adhesion between cells and extracellular matrix in a Ca2+-independent manner. N-CAM is widely expressed during development and plays a crucial role in cell division, migration, and differentiation, but its expression is restricted in adults. The distribution of N-CAM immunoreactivity in adult rat tissues was investigated in the present study. N-CAM immunoreactivity was present in the nervous system in the molecular layer of the cerebellum, ependymal cells surrounding the central canal, axons of the white matter, and in Lamina X of the gray matter of the spinal cord. N-CAM immunoreactivity also was found in autonomic nerves. In the digestive system, N-CAM immunoreactivity was found in the stratified squamous epithelium and nerve plexus of the esophagus, glandular cells of the stomach and pylorus, lamina propria, and epithelium of the villi of the duodenum, jejunum, and ileum. N-CAM immunoreactivity was demonstrated in the secretory cells of the adenohypophysis, islets of Langerhans, and acinar cells of the exocrine pancreas. Alveolar cells of the lung were also N-CAM immunoreactive. In the urinary system, N-CAM immunoreactivity was seen in the proximal convoluted tubules of the kidney. In the male reproductive system, N-CAM immunoreactivity was demonstrated in the nerve plexus around the urethral epithelium and in the nerve fibers around the smooth muscle cells of the corpus cavernosum penis. In the visual system, N-CAM immunoreactivity was seen in the epithelial cells of the corpus ciliaris. Cornea and lens epithelium also showed positive immunoreactivity. Our results suggest that cells in many tissues and organs of the adult rat synthesize N-CAM.

The anatomy of lamina pretrachealis fasciae cervicalis.

The definitions concerning the fascia pretrachealis is either contradictory or insufficient in anatomy textbooks. The fascia pretracheatis is clinically important in the procedure of tracheostomy, mediastinascopy and also in tracheal and bronchial trauma. The anatomy of the fascia pretrachealis (extension, relation and the attachments) was reexamined using cadaveric preparations and the clinical value of the fascia is reinforced. The fascia pretrachealis is attached to the upper brim and to the oblique line of the thyroid cartilage and continued its course on the anterior surface of the trachea and fused with the advantitia of arch of the aorta, posterior aspect of pulmonary artery and the pericardium. Laterally it is attached to the cartilagenous part of the trachea. Also contraversial literature concerning description of the fascia pretrachealis has been evaluated.