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Angiogenesis plays a key role in bone formation and maintenance. Bone formation has been reported to initiate in the concavities rather than the convexities in a hydroxyapatite substratum and the implant threads of dental implants. The aim of the present study was to evaluate the number of the blood vessels inside the concavities and around the convexities of the threads of implants in a rabbit tibia model. A total of 32 thread-shaped implants blasted with apatitic calcium phosphate (TCP/HA blend) (Resorbable Blast Texturing, RBT) (Maestro, BioHorizons(®), Birmingham, AL, USA) were inserted in 8 rabbits. Each rabbit received 4 implants, 2 in the right and 2 in left tibia. Implants were retrieved after 1, 2, 4, and 8 weeks and treated to obtain thin ground sections. Statistically significant differences were found in the number of vessels that had formed in the concavities rather than the convexities of the implants after 1 (p = 0.000), and 2 weeks (p = 0.000), whilst no significant differences after 4 (p = 0.546) and 8 weeks (p = 0.275) were detected. The present results supported the hypothesis that blood vessel formation was stimulated by the presence of the concavities, which may provide a suitable environment in which mechanical forces, concentrations and gradients of chemotactic molecules and blood clot retention may all drive vascular and bone cell migration.

Objective To perform an immunohistochemical evaluation of microvessel density (MVD), vascular endothelial growth factor (VEGF) and nitric oxide synthase (NOS) expression in mandibular ridge regeneration using an equine-derived bone substitute. Materials and methods Ridge augmentations were performed in six patients using equine bone. After a six-month healing period, four bone cores from no augmented (control) and four from augmented sites (test) were retrieved and processed for immunohistochemistry. Results Statistically, significant differences were found in the MVD expression between control and test sites (p Conclusions Within the limits of the small sample size, the present study indicated that equine bone is able to support neovascularization when used as a grafting material.

The aim of the present study was to perform a comparative histological and immunohistochemical evaluation of microvessel density, vascular endothelial growth factor, and nitric oxide synthase (NOS) expression in sinus augmentation using autologous bone alone or in combination with equine bone.Sixteen patients underwent sinus augmentation with autologous bone and a 50:50 mixture of autologous and equine bone. Control cores were harvested from preexisting nonaugmented bone under the sinus floor. The specimens were processed for immunohistochemistry.The greatest values of microvessel density were found in the sites grafted with autologous bone with significant differences between control versus autologous group (P0.01) and control versus autologous + equine group (P0.01). The higher and lower intensities of vascular endothelial growth factor and NOS3 expression were prevalent in the sites grafted with autologous bone with significant differences with the controls (P0.05). No significant differences (P0.05) were found in the NOS1 expression among the groups.The results obtained showed that the mixture of autologous and equine bone was biocompatible, and its use was associated with new blood vessels ingrowth during healing, which has been found to be extremely important for bone formation.

This immunohistochemical study evaluated the inflammatory infiltrate with its cluster differentiation markers (CD 4, CD 8, CD 20, and CD 68) in aggressive and chronic periodontitis gingival tissues in order to identify the specific cell distribution. Twenty-seven human gingival biopsies were obtained and analyzed. Fourteen patients were suffering from chronic periodontitis and six from aggressive periodontitis; seven patients with healthy gingiva were included as the control group. The specimens were immunohistochemically stained for anti-CD 4 (T helper cells), anti-CD 8 (T cytotoxic/suppressor), anti CD-20 (B plasma cells) and anti CD-68 (macrophages). Chronic periodontitis samples were mainly dominated by CD 4 and CD 8+ cells. On the contrary, in aggressive periodontitis patients all four cell types (CD 4, CD 8, CD 20 and CD 68 + cells, respectively) were remarkably increased. CD 20+ cells were significantly (p < 0.05) more prevalent in aggressive versus chronic periodontitis. The control samples expressed lower CD 4, CD 8, CD 20 and CD 68+ cells confirming a none inflammatory state. The present study demonstrates prevalence for CD 20+ cells in aggressive periodontitis lesions. However, further studies need to be performed to confirm and identify a clear pattern of inflammatory cells and hereafter the mechanisms sustaining the disease.

Objective: The aim of this study was to assess vascular endothelial growth factor (VEGF) expression and microvessel density (MVD) in maxillary sinus augmentation with autogenous bone and different graft materials for evaluating their angiogenic potential. Methods: Biopsies were harvested 10 months after sinus augmentation with a combination of autogenous bone and different graft materials: hydroxyapatite (HA, n = 6 patients), demineralized freeze-dried bone allograft (DFDBA, n = 5 patients), calcium phosphate (CP, n = 5 patients), Ricinus communis polymer (n = 5 patients) and control group – autogenous bone only (n = 13 patients). Results: In all the samples, higher intensities of VEGF expression were prevalent in the newly formed bone, while lower intensities of VEGF expression were predominant in the areas of mature bone. The highest intensity of VEGF expression in the newly formed bone was expressed by HA (P

The aim of the present study was to compare the expression of transforming growth factor (TGF) beta1 in ameloblastomas (AMs) with different risk of recurrence by immunohistochemistry. A total of 29 cases of AMs were evaluated. The tumors were divided into 2 groups: group A (10 cases) composed of unicystic and peripheral AMs, associated with a low risk of recurrence, and group B (19 cases) composed of solid AMs, associated with a high risk of recurrence. Statistical evaluation showed significant differences between groups A and B lesions, with higher values of positivity for TGF-beta1 in stromal cells in group B tumors (P = 0.0308). No statistically significant differences of immunoreactivity were found in vessels and odontogenic epithelium between the 2 groups. The increased TGF-beta1 expression in tumors with a high risk of recurrence could be explained with the fact that although TGF-beta acts as a potent tumor suppressor in the early stages of tumor progression, later it seems to enhance the invasive phenotype of the tumor.

Introduction: Recently the term Keratocystic Odontogenic Tumor (KCOT) has been recommended for Odontogenic Keratocysts (OKC) to address the neoplastic nature of the lesion compared to radicular and dentigerous cysts. S100 are calcium-binding proteins involved in cell differentiation and inflammation, with a potential role in neoplastic transformation. Aim: The aim of this study was to evaluate whether S100A8 protein expression is different in KCOT compared to radicular cysts (RC) and dentigerous cysts (DC). Methods: A total of 84 consecutive odontogenic cysts, 34 RC, 25 DC, and 25 KCOT, were analyzed in this study. Results: Epithelial cells in KCOT cases were not immunoreactive for S100A8 except focally in cases associated with inflammation, while RC cases showed a variable positivity of all the epithelial layers from the basal to the superficial in 19/34 cases and DC cases showed a weak positivity of the intermediate and superficial layers in 7/25 cases. Conclusion: The lack of S100A8 protein expression seems to be observed more frequently in KCOT compared to RC and DC. This difference might be related to their neoplastic nature and a potential aggressive biological behavior for odontogenic cystic lesions.

The aim of this study is to evaluate whether hyperoxia and age can influence the expression of vascular endothelial growth factor (VEGF) in the salivary glands of rats. Our study was carried out on four groups of male Wistar rats (total 24 rats). One group was composed of 6 young rats (3 months old); another group by 6 old rats (24 months old). The rats belonging to the first two groups were exposed to hyperoxia (98–100% O2) for a period of 60–65 hours in a large Plexiglas chamber; the other two groups were control groups. The rats were then anaesthetized with Nembutal (30 mg/kg) intraperitoneally and then killed. The submandibular glands were removed and processed for immunohistochemical analysis of VEGF. The exposure to hyperoxia decreased salivary gland VEGF expression in rats. The tissues analyzed (nervous tissue, vascular endothelium, myoepithelial cells, ductal endothelium, mucinous glands) always expressed VEGF, thus demonstrating that not only vascular endothelial cells, but also the other elements evaluated, have a role in the neoangiogenesis. Only in the serous glands, in both normoxic and hyperoxic young and old rats, was the VEGF expression constantly negative and it did not influence the neoangiogenesis. The vascular growth is a fundamental part of normal salivary gland development, so we speculated that strategies aimed at preservation or promotion of salivary gland VEGF expression may mitigate or attenuate hyperoxia-induced gland microvascular injury. Further studies specifically aimed at investigating these prospects are warranted.

The aim of the present study in humans was to conduct a comparative immunohistochemical evaluation of vascular endothelial growth factor (VEGF) and nitric oxide synthase (NOS) expression, inflammatory infiltrate, proliferative activity expression, and microvessel density (MVD) in peri-implant soft tissues of titanium and zirconium oxide healing caps.Five patients, three men and two women (aged 30 to 66 years; mean: 49 years), participated in this study. All patients received dental implants that were 3.8 mm in diameter and 11 mm in length. All implants were left to heal in a non-submerged (single-stage) mode. Healing caps (3.8 mm in diameter and 3.0 mm in height) were inserted in all implants. Half of the implants were supplied with standard, prefabricated caps of commercially pure titanium, whereas the other half were provided with test zirconium oxide caps. After a 6-month healing period, a gingival biopsy was performed with a circular scalpel (5.5 mm in diameter) around the healing caps of both groups, without unscrewing or removing the healing caps. The dimensions of the gingival biopsies were 1.7 mm (5.5-3.8 mm) in thickness and 3 mm in height.Statistically significant differences were found in the microvessel density between titanium and zirconium oxide healing caps and group II (Por = 0.0001). Statistically significant differences were likewise found in the low and high intensities of NOS1, NOS3, and VEGF (Por = 0.0001). In conclusion, the high intensity of NOS1, NOS3, and VEGF were mostly expressed in the titanium group, whereas the low intensity of NOS1, NOS3, and VEGF were mostly expressed in the zirconium oxide group.In our specimens, the inflammatory infiltrate was mostly present in the titanium specimens. Their extension was much larger than that of the zirconium oxide specimens. Higher values of MVD were observed in the titanium specimens (29.1 versus 15.8). In addition, a higher expression of VEGF intensity was observed in the peri-implant tissues of titanium healing caps, whereas predominantly lower expressions of VEGF intensity were noted around the zirconium oxide healing caps. The Ki-67 expression was higher in the titanium specimens. All these data revealed that the tissues around titanium healing caps underwent a higher rate of inflammation-associated processes, most probably correlated to the higher inflammation processes observed in these tissues. A higher intensity expression of NOS1 and NOS3 was recorded in the tissues around titanium, whereas, on the contrary, a lower intensity of expression was found in the tissues around zirconium oxide specimens. These latter data indicate that the higher expression of these two mediators could be correlated to the higher amount of bacteria present around the titanium samples.

The aim of this study was to investigate the differences that are present between apoptosis in symptomatic (with symptoms of cerebral ischemic attack) and asymptomatic carotid atherosclerotic plaques. The apoptotic process in macrophages and smooth muscle cells was evaluated. Cellular markers and products of immune cells in symptomatic and asymptomatic atherosclerotic plaque and endoarterectomy specimen were analyzed by immunohistochemistry. No statistically significant differences were present regarding the mean SMC actin-positive area. Using double staining of α-smooth muscle actin and TUNEL techniques, the number of smooth muscle cells in apoptosis was statistically higher in symptomatic plaque as compared with asymptomatic plaque. Statistically significant differences (p=0.009) were also found in the CD45-positive cells in the inflammatory infiltrate. The CD68-positive macrophages showed statistically significant differences (p=0.0001). Similarly, the double staining with CD68 and TUNEL revealed that apoptotic macrophages were mainly present in asymptomatic plaques rather than symptomatic plaques. Statistically significant differences (p

Background- The clinical benefits of statins are attributed to changes in plaque composition that lead to reduced metalloproteinase (MMP) activity and plaque stabilization. However, the molecular mechanism of this effect is unclear. Recently, we demonstrated enhanced expression of isoforms of inducible cyclooxygenase (COX) and PGE synthase (COX-2/mPGES) in human symptomatic plaque and provided evidence that this is associated with MMP-induced plaque rupture. The aim of this study was to characterize the effect of simvastatin on inflammatory infiltration and the expression of COX-2/mPGES and MMPs in human carotid plaques. Methods and Results----Seventy patients with symptomatic carotid artery stenosis were randomized to the American Heart Association Step 1 diet plus simvastatin (40 mg/d) or the American Heart Association Step 1 diet alone for 4 months before endarterectomy. Plaques were subjected to analysis of COX- 1, COX-2, mPGES, MMP-2 and MMP-9, lipid and oxidized LDL (oxLDL) content, and collagen content by immunocytochemistry, Western blot, and reverse transcription-polymerase chain reaction, whereas zymography was used to detect MMP activity. Immunocytochemistry was also used to identify CD68+ macrophages, CD3+ T-lymphocytes, smooth muscle cells (SMCs), and HLA-DR+ inflammatory cells. Plaques from the simvastatin group had fewer (P

Fractal dimension (FD) in tissue specimens from patients with oral squamous cell carcinoma (OSCC) was evaluated. FD values in different stages of OSCC, and the correlations with clinicopathological variables and patient survival were investigated. Histological sections from OSCC and control non-neoplastic mucosa specimens were stained with hematoxylin-eosin for pathological analysis and with Feulgen for nuclear evaluation. FD in OSCC groups vs. controls revealed statistically significant differences (P

The human DNA mismatch repair (hMMR) system plays an important role in reducing mutation and maintaining genomic stability. The MMR system in human cells is composed of at least six genes (hMSH2, hMLH1, hMSH3, hPMS1, hPMS2 and GTBP/hMSH6). In particular, hMSH2 and hMLH1 are expressed in human cells that are undergoing rapid renewal; their reduced expresion has been reported in several tumors. We examined the protein expression pattern of hMSH2 and hMLH1 by immunohistochemistry in 25 ameloblastomas. All ameloblastomas expressed hMSH2 and hMLH1 proteins in the outer layer of epithelial cells. The localization of the staining was exclusively nuclear. These data suggest that the development and progression of these tumors do not depend on a defect in the hMMR system.

An experimental murine model was studied to evaluate the orogastrointestinal colonization of Helicobacter pylori and the animal-to-animal transmission. Balb/C mice were infected with H. pylori and housed with uninoculated mice in cages with and without a grate on the floor. Mice were killed after 7, 14, 30, and 45 days, and samples from the esophagus, stomach, small intestine, colon, and rectum were analyzed for H. pylori by PCR and immunohistochemistry and for histological changes. Bacterial colonization was assessed also by culture from stomach samples. H. pylori was cultured by stomach samples of infected mice at 7, 14, and 30 days. Using PCR and immunohistochemistry, H. pylori was detected in inoculated and uninoculated mice in all areas examined, with an high percentage of positive samples in the esophagus and stomach. Moreover transmission was detected, without differences, regardless of whether mice were housed with or without a grate on the floor, supporting an orooral animal transmission.

Bone tissue is characterized by a constant turnover in response to mechanical stimuli, and osteocytes play an essential role in bone mechanical adaptation. However, little to no information has been published regarding osteocyte density as a function of implantation time in vivo. The aim of this retrospective histological study was to evaluate the osteocyte density of the peri-implant bone in implants retrieved because of different reasons in a time period from 4 weeks to 27 years. A total of 18 samples were included in the present study. Specimens were divided into 3 groups depending on the loading history of the implants: loading between 4 weeks and 7 months (group 1); loading between 1 and 5 years (group 2); loading between 14 and 27 years (group 3). All the samples were histologically evaluated and osteocyte density was obtained using the ratio of the number of osteocytes to the bone-area (mm(2) ). The osteocyte density values significantly increased in the Group 2 (1-5 years) compared with Group 1 (4 weeks-7 months), and significantly decreased in the Group 3 (14-27 years) compared to Group 2. No significant differences were detected between Group 1 and Group 3. The decrease in osteocyte density observed in samples that were in vivo for long periods of time under loading is possibly because of the fact that once the bone structure is well aligned and biomechanically competent, a lower number of osteocytes are necessary to keep the tissue homeostasis under loading.

The cause of sarcoidosis is unknown, although mycobacteria have been implicated. We examined sarcoid tissues for human herpesvirus 8 (HHV-8) in addition to mycobacterial genomic sequences.Biopsy samples from 17 patients with sarcoidosis were studied (eight transbronchial, 27 lymph node, two skin, and two oral mucosa). We used tissues (n = 137) from 96 patients without sarcoidosis as negative controls. A nested PCR was applied to amplify a segment of open reading frame (ORF) 26 of the HHV-8 genome, and a heminested PCR was to amplify a segment of ORF 25 of HHV-8 and of the 16 S rRNA gene of mycobacteria. Differences in base sequences of the amplified fragments were resolved with single-strand conformation polymorphism and dideoxy sequencing.HHV-8 ORF 26 DNA was detected in significantly higher proportions of sarcoid than of non-sarcoid tissue samples from lung (8/8 vs 0/54; p0.0001), lymph nodes (26/27 vs 6/29; p0.0001), skin (2/2 vs 0/17; p = 0.006), and oral tissues (2/2 vs 1/13; p = 0.029). 31 (82%) of the 38 ORF 26 DNA-positive sarcoid specimens were also positive for ORF 25 DNA. For mycobacteria-like 16 S rRNA DNA, the proportion positive was significantly higher in sarcoid than non-sarcoid tissues for lymph node samples (11/27 vs 2/29; p = 0.003) but not for other tissues (lung 3/8 vs 22/54; skin 2/2 vs 15/17; and oral tissues 1/2 vs 0/13). Overall, the prevalence of HHV-8 ORF 26 sequences was higher in sarcoid tissues than in non-sarcoid tissues (p0.0001). When patients whose tissues were included in a masked phase of the study were treated as units of analysis, eight of eight sarcoidosis patients were positive for HHV-8 ORF 26 DNA, compared with three of 56 control patients (p0.0001); for mycobacteria-like sequences, three of eight sarcoidosis patients were positive, compared with four of 56 controls (p = 0.0464). The HHV-8 ORF 26 sequences, ten of which were unique, could be segregated into four groups according to peptide motifs. In seven of nine patients from whom biopsy samples were taken from various sites, different sequences were recovered. The mycobacterial sequences amplified from sarcoid tissues were also varied, but none was homologous to those of known species.Variant HHV-8 DNA sequences are found in a wide range of sarcoid but not non-sarcoid tissues. Mycobacteria-like 16 S rRNA sequences are more frequently present in sarcoid lymph nodes and not in other tissue types, but do not indicate infection by a particular mycobacterial species.

A 210-bp DNA segment specific to the human herpesvirus 8 (HHV-8) genome was amplified by nested polymerase chain reaction from 10 of 14 archived oral biopsy samples of HIV-positive patients in London who had no evidence of oral Kaposi's sarcoma (KS). Various oral sites were represented. Oral tissues from 20 general dental patients not known to be HIV-infected were negative. When DNA sequences of these products were compared with sequences derived from 5 oral KS tissues of AIDS patients in London and 10 skin biopsies of Italian patients with Mediterranean KS (total number of positive tissues = 25), 11 were found to be unique. DNA and predicted peptide motifs of these sequences were also different from those in 28 of 36 HHV-8-positive lesions previously reported from American and African patients. HHV-8 is tropic for the oral mucosa of HIV-infected persons, and HHV-8 variants, though diverse, may be geographically restricted.

Oral lichen planus (OLP) is a common chronic inflammatory disorder sometimes associated with hepatitis C virus (HCV) infection. An increased prevalence of autoimmune markers has been reported in patients with HCV infection. The aim of the present study was to determine, by conventional indirect immunofluorescence, the nature and frequency of circulating antibodies to epithelial antigens in the sera of HCV-positive patients who also have OLP. The study comprised four groups: 14 patients with OLP and HCV infection. 14 HCV-seronegative patients with OLP, 21 HCV-seropositive patients without OLP and 18 healthy controls. We found a significant association between the concomitance of OLP and HCV infection and the presence of such antibodies. It is concluded that some patients with HCV-associated OLP may have circulating antibodies to epithelial antigens, although their precise aetiological role in the development of this disease in HCV infection remains unknown.

PURPOSE This human study sought to compare, from an immunohistochemical point of view, matrix metalloproteinases (MMPs) 2, 3, 8, 9, and 13 in the soft tissues around titanium and zirconium oxide healing caps. MATERIALS AND METHODS Five patients participated in this study. All patients received 3.8 × 11-mm dental implants, which were left to heal in a nonsubmerged (single-stage) mode. Healing caps (3.8 mm in diameter and 3.0 mm in height) were inserted in all implants. Half of the implants were randomly supplied with standard, prefabricated caps of commercially pure titanium (control), while the other half were randomly provided with zirconium oxide caps (test). After a 6-month healing period, gingival biopsy specimens were obtained with a circular scalpel around the healing caps of both groups, without unscrewing or removing the healing caps, and the samples underwent immunohistochemical processing for MMPs 2, 3, 8, 9, and 13. RESULTS Statistically significant differences were found in the values of MMP-8 in the cells of the inflammatory infiltrate, with higher values for the titanium samples. Statistically significantly higher values were found, also in the titanium samples, for MMP-9 in the endothelial cells of the blood vessels. No statistically significant differences were found for any other MMPs. CONCLUSIONS The present results showed that the soft tissues around titanium healing caps underwent a higher rate of restorative processes, most probably correlated to the MMP levels observed in the tissues.

Introduction Calcium sulfate (CaS) is a simple, biocompatible material with a long history of safe use in different fields of medicine. CaS is a rapidly resorbing material that leaves behind a calcium phosphate lattice, which promotes bone regeneration and hemostasis. The aim of this study was a clinical evaluation of the hemostatic effect of CaS hemi-hydrate (CaSO4), commonly known as plaster of Paris, in endodontic surgery. Methods Twenty-four patients with 31 periradicular lesions were enrolled in this study. The apical roots were exposed, and the bleeding would have made it difficult to correctly fill the root-end cavities. To avoid such an inconvenience, the teeth were divided into 3 groups. Hemostasis was attempted by using CaS in 11 teeth (group I), gauze tamponade in another 10 teeth (group II), or 20% ferric sulfate in the last 10 teeth (group III). Results Control of the bleeding was achieved in all teeth of group I, whereas in group II adequate hemostasis was achieved in 3 of 10 cases and in group III in 6 of 10 cases. Conclusions The use of CaS completely eliminated the bleeding, with a very good level of hemostasis.

The route of Helicobacter pylori transmission remains unclear and the currently suggested route is person-to-person transfer by faecal-oral and oral-oral mode. The aim of this study was to verify the presence of H. pylori in esophagus and saliva of humans. Saliva samples, mucosal biopsies from esophagus, gastric antrum and fundus were collected from 19 patients with positive Urea Breath Test (UBT). Gastric biopsies were used for H. pylori colture and antimicrobial susceptibility tests whereas saliva samples were collected to detect H. pylori with a Nested-PCR targeting 16S rRNA gene as well as esophagus biopsies which were also investigated with immunohistochemical staining. Helicobacter pylori was isolated in 18 patients both in gastric antrum and fundus. The molecular analysis, confirmed by comparative sequences evaluation, gave positive results in all saliva and esophageal samples whereas the immunohistochemistry revealed the presence of H. pylori in 15.8% (3/19) of the esophagus samples. Our data suggest that saliva and esophagus may be considered reservoirs for H. pylori in humans and emphasize the need to use more susceptible techniques for H. pylori detection, in particular in over-crowded sites. Identification of the transmission route of H. pylori is crucial in developing an effective plan of surveillance by finding new means of disease management.

p53 protein seems to be related to the suppression of cell proliferation. p53-positive tissues seem to have a higher proliferative activity than p53-negative ones. Odontogenic keratocyst (OKC) has a different behavior from other types of cysts because it is more aggressive, with a tendency to recurrence. Twenty-two dentigerous cysts, 24 radicular cysts, and 20 OKCs were used in the present study. Two dentigerous cysts (9.1%), 2 radicular cysts (8.3%), and 9 OKCs (45%) expressed the p53 protein. The differences between the three groups were statistically significant (p = 0.003). In 10 cases of OKCs epithelial dysplasia was found. One of the 10 OKCs without dysplasia and 8 of the 10 OKCs with dysplasia were p53-positive: the difference between the two groups was statistically significant (p = 0.007). The overexpression of p53 protein was not on the other hand correlated with the occurrence of multiple, bilateral, and recurrent OKCs. Moreover the distribution of p53-positive cells was parabasal in contrast with other types of cysts. These qualitative and quantitative differences in proliferative activity in OKCs seem to point to an alteration in cell cycle control.

In periodontal tissues, angiogenesis seems to be important for the maintenance of healthy tissues and in periodontal diseases. Angiogenesis is regulated through a complex interplay of molecular signals mediated by growth factors involving extracellular matrix remodeling, endothelial cell migration and proliferation, capillary differentiation and anastomosis. However, the influence of angiogenesis in the development, progression, and healing of periodontal lesions is currently under investigation. This immunohistochemical study evaluates the expression of vascular endothelial growth factor (VEGF), microvessel density (MVD), nitric oxide synthase (NOS) 1 and 3, and Ki-67 in gingival tissues of patients with aggressive and chronic periodontitis.Twenty-seven human gingival biopsies were taken from patients with chronic periodontitis (n = 14 patients), generalized aggressive periodontitis (n = 6 patients), and healthy periodontia (n = 7 patients). The specimens were immunohistochemically stained for VEGF, MVD, NOS 1 and 3, and Ki-67.The levels of VEGF, MVD, NOS 1 and 3, and Ki-67 were found to be significantly different among groups (P0.001). Patients with aggressive periodontitis had higher levels of these markers compared to those in patients with chronic periodontitis and healthy patients.The analysis demonstrates a higher expression of all immunologic markers particularly in subjects with aggressive periodontitis. In summary, the data from this pilot investigation suggests that VEGF is an important factor in the pathogenesis of the aggressive and chronic forms of periodontitis.

Background: The aim of the present study is a comparative evaluation of inflammatory infiltrate, microvessel density, vascular endothelial growth factor, nitric oxide synthase, and proliferative activity in soft tissues below intraorally welded titanium bars. Methods: Twenty-two patients participated in this study. All patients carried immediately loaded one-stage titanium implants splinted with intraorally welded titanium bars. Each patient underwent two gingival biopsies, a control biopsy harvested from an area of mucosa 5 mm away from the titanium bar and a test biopsy from the mucosa below the titanium bar, which were histologically and immunohistochemically processed. Results: No fractures or radiographically detectable alterations of the welded frameworks were present. In all the cases examined, the average of the modified plaque index was 1, no suppuration or bleeding on probing was present, and probing depth was £3 mm. However, the immunohistochemical analysis revealed some differences. The inflammatory infiltrate was mostly present in test sites and its extension was much larger than in control sites. Statistically significant differences were found in microvessel density and Ki-67 expression among control and test groups (P

The oral cavity, and particularly the gingival mucosa, is continuously exposed to numerous food and bacterial plaque antigens, though evident immunologic reactions are uncommon. It is therefore possible that the mucosal associated lymphoid tissue (MALT) of this region is preferentially biased towards unresponsiveness, rather than immune cell activation. The distribution and phenotype of immune cells in normal human gingiva were examined. Their distribution varied, and high and low cellularity areas could be distinguished in the same specimen. The number of CD3 positive (CD3+) T lymphocytes was more than thrice higher in a high cellularity area. In both types of area, intraepithelial T lymphocytes were not activated. Moreover, they showed chromatin condensation and cell shrinkage characteristic of apoptosis. In the stroma of high cellularity areas, foci of cell activation and numerous B cells were present, suggesting a localized active immune response. The vast majority of intraepithelial and stromal T lymphocytes expressed the "memory" CD45RO+ phenotype. The absence of an immune response within the epithelium and the localized response in the stroma (probably due to the binding of memory T cells to antigens in a low affinity, cross-reactive fashion) may be a part of a protective mechanism against indiscriminate stimulation by a multitude of external antigens.

The objective of the present study was to evaluate vascular endothelial growth factor (VEGF) expression in different types of odontogenic cysts. A total of 25 parakeratotic odontogenic keratocysts (POKCs), 16 orthokeratotic odontogenic keratocysts (OOKCs), and 28 follicular cysts (FCs) were evaluated semiquantitatively for immunohistochemical analysis of VEGF in epithelial cells, endothelial cells of blood vessels, inflammatory cells and focally stromal cells. A significant different expression of VEGF in all cell components was found in keratocysts compared to FCs. The POKCs (80%) and OOKCs (68%) showed more than 50% VEGF positive epithelial cells, whereas the majority of FCs (71%) were either negative in the epithelium or showed less than 10% positive cells. Similarly, the POKCs (88%) and OOKCs (68%) showed more than 50% positive endothelial cells, whereas the FCs (75%) were either negative or showed less than 10% VEGF positive endothelial cells. The highest percentage of cases with score 2 positivity in the stromal cells was observed in POKCs (68%); OOKCs showed a score 2 positivity in 44%, score 1 in 31% and score 0 in 25%, whereas 68% of FCs showed a score 0, 25% a score 1 and only 7% of cases showed a score 2. No statistically significant differences were observed between POKCs and OOKCs in VEGF expression in the epithelial and endothelial cells, whereas the positivity score in stromal cells was significantly higher in POKCs compared to OOKCs. The present results can support the hypothesis that angiogenesis is an active mechanism in the invasive growth of the OKC.

A close spatial correlation has been described between the roughness of intraoral materials and the rate of bacterial colonisation. The aim of the present study in man was to conduct a comparative immunohistochemical evaluation of the inflammatory infiltrate, microvessel density, the nitric oxide synthases 1 and 3 and the vascular endothelial growth factor expression, the proliferative activity, and the B and T lymphocyte and histiocyte positivity in the peri-implant soft tissues around machined and acid-etched titanium healing caps. Ten patients participated in this study. The patients were enrolled consecutively. All patients received dental implants left to heal in a non-submerged mode. Healing caps were inserted in all implants. Half of the implants were supplied randomly with machined caps of titanium (control), while the other half were provided randomly with acid-etched titanium caps (test). After a 6-month healing period, a gingival biopsy was performed with a circular scalpel around the healing caps of both groups. The inflammatory infiltrate was mostly present in test specimens. Their extension was much larger than that of the control samples. A higher number of T and B lymphocytes were observed in test specimens. Higher values of microvessel density and a higher expression of vascular endothelial growth factor intensity were observed in the test samples. Furthermore, the Ki-67, NOS1 and NOS3 expression was significantly higher in the test specimens. All these results showed that the tissues around test healing caps underwent a higher rate of restorative processes, most probably correlated to the higher inflammation processes observed in these tissues.

Neuroglobin (Ngb) is a member of the vertebrate globin family expressed particularly in the brain and in the retina. Ngb is concentrated in the mitochondria-containing areas of neurons, and its distribution is correlated with oxygen consumption rates. Previously we have shown that Ngb is expressed in carotid body (CB) tissues. Considering that hypoxia and aging may be linked through a series of adaptive and protective mechanisms (e.g. reduction in mitochondrial numbers), we investigate the role of Ngb during aging and hypoxia. Two groups of six rats (age-matched 3 and 24 months old) were kept in room air as a control groups, the others two groups were kept in a Plexiglas chamber for 12 days in chronic hypoxia (10–12% inspired oxygen). The presence of Ngb in the CB tissue was detected by immunohistochemistry using a polyclonal antibody. Ngb immunoreactivity was significantly higher in CB tissues from young rats exposed to chronic intermittent hypoxia, whereas CB tissues from old rats did not show any significant increase in Ngb levels after hypoxia. Similar to hemoglobin, Ngb may act as a respiratory protein by reversibly binding gaseous ligands NO and O2 and could act as a NO scavenger and participate in detoxification of Reactive Oxygen Species (ROS) generated under hypoxic conditions.

Background: In the case of localized ridge atrophy, a ridge augmentation procedure, with the use of bone substitutes and barrier membranes, may then be necessary. Purpose: The aim of the present study was a clinical, histological, and immunohistochemical evaluation of an equine spongy bone in alveolar ridge augmentation procedures. Materials and Methods: Five patients showing horizontal mandibular ridge defects participated in this study. A ridge augmentation was performed through an onlay apposition of equine bone covered by a titanium-reinforced membrane. After 6 months of healing, five bone cores from nonaugmented sites (control) and five from augmented sites (test) were retrieved. Results: In test sites, no postoperative complications occurred. Horizontal bone width increased from 2 4t o37 mm. In control sites, the newly formed bone represented 33%, and in test sites, 35% of the total area. The mean value of the microvessel density was 25.6 +/– 3.425 per mm 2 in controls, while 33.3 +/– 2.5 vessels per mm 2 in the test sites were found (p < .05). Both groups showed a high intensity (++) of vascular endothelial growth factor expression in the newly formed bone, while a low intensity (+) was found in the mature bone. Conclusion: Equine bone appeared to be biocompatible and to be associated with new vessel ingrowth. Within the limits of the small sample size, the present study indicated that equine bone could be used in mandibular ridge augmentations.

Despite advances in biological and molecular characteristics, the prognosis of oral squamous cell carcinomas is still very unfavourable and is based on the classical clinicopathological parameters. However, tumors with similar clinicopathological characteristics may differ dramatically in their clinical outcome. Thus, the identification of novel prognostic factors is necessary to improve prognostic and therapeutic approaches. Transforming growth factor-beta1 (TGF-beta1) is a potent growth inhibitor of epithelial cell proliferation, thus, inactivation of TGF-beta1 signalling may play a role in cancer. The expression levels of TGF-beta1 and its type I and type II receptors (TbetaRI and TbetaRII) were assessed by immunohistochemical and Western blot analyses in 22 oral squamous cell carcinoma lesions, in their normal adjacent mucosa and in the squamous carcinoma cell lines FaDu and CAL27. Immunohistochemistry on 22 oral carcinomas and case-matched normal oral mucosae demonstrated that TGF-beta1, TbetaRI, and TbetaRII were intensively and homogeneously expressed in all normal epithelia. In contrast, TGF-beta1 and its receptors were significantly reduced in poorly (G3) differentiated tumors as compared to moderately (G2) and well differentiated (G1) lesions (p=2.8 x 10(-3), p=1.3 x 10(-3), p=2.8 x 10(-3) and p=1.3 x 10(-3), respectively). The progressive reduction of the expression levels was confirmed by Western blotting. The oral squamous carcinoma cell lines Cal27 and FaDu demonstrated a reduced and a lack of TbetaRI expression, respectively. A significant decrease of TbetaRII expression, as compared to Cal27 cells, was shown in FaDu cells. Thus, the decreased expression of TbetaRII combined with the absence of TbetaRI could account for the resistance of FaDu cells to the growth-inhibiting effect of TGF-beta1. TGF-beta1 and TGF-beta1 receptor expression significantly decreased as tumors became less differentiated and thus more aggressive, suggesting a functional role of these molecules in oral tumor progression.

Purpose This study was an immunohistochemical evaluation of microvessel density (MVD) in sinus augmentation procedures with autologous bone and anorganic bone (Bio-Oss). Materials and methods Twenty-four patients (14 men and 10 women - mean age of 48 years with a range from 34 to 53 years) participated in this study. All the patients presented a maxillary partial unilateral edentulism involving the premolar/molar areas, with a residual alveolar ridge height of about 4 to 5 mm. Twelve patients received sinus augmentation procedures with 100% autologous bone; 100% Bio-Oss was used in the other 12 patients. Endosseous implants were inserted after a mean period of 3 months. As control, the portions of preexisting subantral bone were used. The mean value of the MVD in control bone was 23.4 +/- 1.3. The mean value of the MVD in the sinuses augmented with autologous bone was 29.0 +/- 2.4. The mean value of the MVD in the sinuses augmented with Bio-Oss was 23.8 +/- 2.2. Results The statistical analysis showed that the differences of the MVD between control bone and sinuses augmented with Bio-Oss were not statistically significant (P = 0.52), while the difference of the MVD between sinuses augmented with autologous bone and those augmented with Bio-Oss was statistically significant (P = 0.0008). Conclusions Autologous bone may act not only as a passive filling material in bone defects but may also release osteogenic growth factors; and particles of autologous bone seem to contain vital osteoprogenitor cells.

Maspin (mammary serine protease inhibitor) is a member of the serpin superfamily of protease inhibitors and it has a role as a tumor suppressor. Maspin has been reported to be important in processes relevant to tumor growth and metastasis such as cell invasion, angiogenesis, and apoptosis. A high expression of maspin was correlated with better rates of survival and absence of nodal metastases in head and neck squamous cell carcinoma. In contrast, some studies have shown that maspin overexpression is correlated with a poor prognosis in pancreatic and ovarian cancers and in lung adenocarcinoma. The aim of this study was an immunohistochemical evaluation of the maspin expression in oral squamous cell carcinoma and thus 89 patients were evaluated. Maspin expression in oral squamous cell carcinoma was significantly associated with the tumor differentiation grade (chi test: P = 0.0318) and the lymph node status (chi test: P < 0.005), but not with the tumor stage (chi test: P = 0.666). Metastatic involvement of lymph nodes was observed more frequently in maspin-negative cases than in tumors with more than 5% of positive cells (P = 0.0024). The present results confirm that maspin expression predicts a better prognosis in oral squamous cell carcinoma and that maspin probably plays a role in tumor progression.

Statement of problem Ovate pontics mimic the natural tooth contour and provide an esthetic result however, few studies have evaluated the histological changes in underlying tissues. Purpose The purpose of this pilot study was to histologically evaluate the healing of gingival tissues in contact with provisional ovate pontics after 2 weeks. Material and methods Three patients requiring fixed partial dentures participated in this study. The provisional restorations consisted of a fixed partial denture having 2 ovate pontics: one in acrylic resin (Jet), with an ovate shell made of low-fusing ceramic (Duceram LFC); the other made completely of the same acrylic resin (control). After 2 weeks, biopsies of the gingival tissues beneath the pontics were retrieved for histological examination, and immunohistochemistry for evaluation of the vascular endothelial growth factor (VEGF) was performed. Data were analyzed using descriptive statistics. Results Clinically, all of the ovate pontic–prepared sites showed partial healing. Histologically, the thickness of the entire mucosa was similar in both specimens; however, in some regions, the epithelium presented ulcerations that were generally deeper and more frequent in control sites than in test sites. Immunohistochemical results showed that tissues beneath LFC pontics seemed to be less inflamed since they demonstrated a lower expression of VEGF (10.7 ± .8) compared to those beneath acrylic resin ovate pontics (33.9 ± 2.5). Conclusion This pilot study demonstrated that the placement of provisional LFC ovate pontics may be advantageous for the reparative processes of the underlying tissues.

We performed an immunohistochemical study in a series of ameloblastomas with different histology to explore the existence of a correlation between CD10 immunoreactivity in peritumoral stromal cells and the type of ameloblastoma with a high risk of local recurrence.A total of 45 ameloblastomas (18 unicystic [UA], 4 peripheral [PA], 23 solid/multicystic [SA]) were evaluated. Cases showing immunoreactivity for CD10 inandor =10% of stromal cells around tumoral epithelial islands, were considered, respectively, negative and positive. Correlations between stromal CD10 expression and histopathologic types with low and high risk of recurrence were evaluated by statistical analysis.SA cases showed a significantly higher percentage of stromal CD10-positive cells than the UA and PA variants. A strong intensity of immunostaining was observed only in SA.Our results suggest that CD10 expression might be associated with stromal invasion in ameloblastoma variants with a high risk of recurrences.

affinities and binds CO (Burmester & Hankeln, 2004).The physiological role of Ngb is not well understood, but it has been proposed that Ngb participates in several processes such as oxygen transport, oxygen storage, and NO detoxification (Burmester and Hankeln, 2004). Ngb as well as hemoglobin is a respiratory proteinthat reversibly binds gaseous ligands (NO and O

Crestal bone loss has been reported to occur around dental implants. Even if the causes of this bone loss are not completely understood, the presence of a microgap between implant and abutment with a possible contamination of the internal portion of the implants has been suggested. The aim of this study was to see if there were differences in the vascular endothelial growth factor (VEGF) expression, microvessel density (MVD), proliferative activity (MIB-1), and inflammatory infiltrate in the soft tissues around implants with screwed and cemented abutments. Sandblasted and acid-etched implants were inserted in the mandibles of 6 Beagle dogs. Ten 3.5- × 10-mm root-form implants were inserted in each mandible. A total of 60 implants (30 with screwed abutments and 30 with cemented abutments) were used. After 12 months, all the bridges were removed and all abutments were checked for mobility. A total of 8 loosened screws (27%) were found in the screwed abutments, whereas no loosening was observed in cemented abutments. A gingival biopsy was performed in 8 implants with cemented abutments, in 8 implants with screwed abutments, and in 8 implants with unscrewed abutments. No statistically significant differences were found in the inflammatory infiltrate and in the MIB-1 among the different groups. No statistically significant difference was found in the MVD between screwed and cemented abutments (P = .2111), whereas there was a statistically significant difference in MVD between screwed and unscrewed abutments (P = .0277) and between cemented and unscrewed abutments (P = .0431). A low intensity of VEGF was prevalent in screwed and in cemented abutments, whereas a high intensity of VEGF was prevalent in unscrewed abutments. These facts could be explained by the effects induced, in the abutments that underwent a screw loosening, by the presence of bacteria inside the hollow portion of the implants or by enhanced reparative processes.

The mechanical properties of bone are greatly influenced by the percentages of organic and mineral constituents. Nevertheless, the information about the mineral content on a microscopic scale in peri-implant bone is scarce. The aim of this work was to analyze the bone mineral density of peri-implant bone under different techniques.Five unloaded titanium dental implants with a micro-structured surface (three XiVE plus and two Frialit 2, DENTSPLY-Friadent, Mannheim, Germany) were retrieved from the mandible of five patients after a 6-month period. scanning electron microscopy with backscattered electron signal (BSE), light microscopy (LM) with a double staining technique, fluorescence microscopy and confocal laser microscopy were used for measuring microscopic mineral content variations in peri-implant bone. Histomorphometry and image intensity (grey level) were evaluated using a software package for image analysis.The low mineral density index (LMDI) for LM was of 29.2+/-3.1 (mean+/-S.D.), while the high mineral density index (HMDI) was of 88.2+/-3.6 (mean+/-S.D.). The one-way ANOVA analysis showed a significant difference (P0.001) among the groups. The pairwise Holm-Sidak test identified the differences among HMDI indexes for both LM and SEM values and also for cross-evaluation of the LMDI and HMDI values. The comparison between LMDI indexes for both SEM and LM did not show any significance. The fluorescence microscopy analysis showed clearly the difference between old (high mineralized) and new (low mineralized) bone tissue near the implant surface. Under confocal laser microscopy the same sections showed the area of bone modelling closest to implant surface.In this study it was found that bone around unloaded implants showed a low mineral density index under all the investigation methods used. It was also found that the conventional LM technique with the double staining method was able to intensely stain the bone area with a low mineral content.

Many oncogenis and tumour suppressor genes found inside normal and pathological cells are fundamental for the processes of development, proliferation and tissue differentiation. The purpose of our study is to show the presence and a possible relationship of the VEGF protein during different phases of the development of human dental germ centers. After cephalometric investigation in 8 orthodontic patients with a mean age of 13 years, (4 females and 4 males), hyperdivergence of the third molars were extracted. The 40 surgical samples were tested with monoclonal human anti-VEGFs antibodies carrying out a semi-quantitative analysis to look for a positive reaction. Reaction for anti-VEGF antibodies was detected in normal embryological tissues and in microvessels near odontogenic cells. During different phases of embryologic development of the dental bud our search showed intracytoplasmatic positive immunoreactions both in the ameloblastic and odontoblastic cells. Additionally, a positive reaction was observed for the VEGF protein in the cells of the stellate reticulum and in those endothelial tissue surrounding the microvessels in all the samples examined.

Oral manifestations are present in about 3-5% of cases of non-Hodgkin's lymphoma (NHL) and oral lesions are only rarely the initial manifestations of NHL. A case is presented of an 80-year-old patient with a NHL of the tongue, without visceral or lymph node involvement. The diagnosis of NHL can be made only by biopsy. The prognosis of NHL seems to be related to the tumour stage, tumour aggressiveness and response to treatment: the oral lesions appear to respond quite well to irradiation.

To test if oxygen sensitive mechanisms are affected by hypoxia, we studied hypoxia inducible factor-1alpha (HIF-1alpha), vascular endothelial growth factor (VEGF) and inducible nitric oxide synthase (iNOS) expression by immunohistochemical analysis in young and old rat carotid bodies (CBs) using hypoxia as a model for modulating aging. Four groups of male age-matched Wistar rats (3 and 24 months) were used. Two groups were kept in room air, and two groups were kept under chronic intermittent hypoxia for 12 days. In aged carotid body and in hypoxia the increased expression of HIF-1alpha, VEGF, iNOS is less evident as compared to the young one. Electron microscopy sections showed a reduced mitochondrial number and area in the aged CBs and during hypoxia. Less responsiveness to hypoxia could be evidenced in the aged rats as compared to the young rats, suggesting an age dependency of the oxygen sensitive mechanisms.

Carotid body (CB) undergoes several morphological, physiological and biochemical changes during aging, hypoxia and hyperoxia (Di Giulioet al., 1998; Lahiriet al., 1990; Lahiri et al.,2000). CB releases several substances like dopamine, acetilcholine, nerpinephrine, erythropoietin, substance P, and it detects pO2 and pCO2 levels in the blood, it regulates ventilation according to oxygen needs and body requirements for homeostasis maintenence (Bunnet al., 1996). The ventilatory response to hypoxia is characterised by increase in volume and ventilatory frequency in relation to the degree of hypoxia. This response is attenuated with aging (Fukuda 1992) and it is related to the age-dependent structures and function modifications (Guenard 1998), including the basal reduction of oxygen requirements (Sohalet.al,1986, Gunnarsson et al, 1996).

Carotid body (CB) is made up neuroepithelial tissue and it exhibits some characteristics which are similar to those of endocrine gland (Lawson, 1980; Paulding et al., 2002). Oxygen chemosensitive mechanisms are present in the carotid body, but the site and precise mechanisms of chemotransduction are still unknown.

Vascular endothelial growth factor (VEGF) is a glycoprotein that has the capability to increase vascular proliferation and permeability. VEGF has been found to be expressed in several different types of tumors, and it may contribute to the progression of malignant tumors. Immunostaining for VEGF and factor VIII was performed in normal healthy pulps and in irreversible pulpitis. In both cases the vessels were always positive for VEGF. Our immunohistochemical data show that the expression of VEGF was strongly positive in the inflammatory infiltrate in irreversible pulpitis. VEGF expression in the stromal cells in healthy pulps ranged from 20 to 100% (with a mean of 68.82), and in irreversible pulpitis ranged from 0 to 100% (with a mean of 62.35%); this difference was statistically significant (p = 0.05). This down-regulation in the stromal cells in irreversible pulpitis could be due to the presence, in a low compliance system such as the dental pulp, of inflammatory infiltrate. VEGF is probably a factor implicated in the etiology and progression of pulpitis. The microvessel density in healthy pulps was 90.00 +/- 27.5, while, in irreversible pulpitis, it was 56.68 +/- 21.15. This difference was statistically significant (p = 0.001). The decrease in microvessel density in irreversible pulpitis could be related to failing vascular function and blood flow decrease.

Background and Aims: Matrix metalloproteinases (MMPs) represent a group of enzymes that regulate cell-matrix composition playing a major role in the inflammatory response. In the present study we evaluated the ability of the MMP inhibitor Batimastat (BB-94) to modify the course of experimental colitis induced in the rat by trinitrobenzensulfonic acid (TNB). Methods: Colitis was induced in 40 rats by intracolonic administration of TNB. Animals were divided into four groups of ten rats each: group 1 received only intracolonic TNB, group 2 received TNB+5 mg/kg intraperitoneal BB-94, group 3 TNB+10 mg/kg BB-94 and group 4 TNB+20 mg/kg BB-94. The MMP inhibitor was administered 30 min before induction of colitis and twice daily until death. Ten rats receiving only intracolonic 0.9% saline served as controls. Animals were killed after seven days; segments of colon were removed and used for histological score of inflammation and myeloperoxidase (MPO) activity. Results: Rats receiving only intracolonic 0.9% saline showed no evidence of colitis. The inflammation score was 0.9, MPO activity 0.235 U/mg. Group 1 (TNB-treated rats) exhibited a high inflammation score (12.4) and MPO activity (0.715 U/mg). Conversely, BB-94-treated rats showed, compared to the TNB group, a significantly lower inflammation score and MPO activity in a dose-dependent fashion. Group 2: inflammatory score 10.1, MPO activity 0.474 (p < 0.05 vs. TNB); group 3: inflammatory score 8.3, MPO activity 0.287 (p < 0.01 vs. TNB); group 4: inflammatory score 5.0, MPO activity 0.256 (p < 0.01 vs. TNB). Conclusions: Treatment with BB-94 has dose-dependent beneficial effects on the inflammatory alterations in rat experimental colitis. Thus, the inhibition of MMPs may represent a novel therapeutic approach for treatment of intestinal inflammation.