Javier Enrique Botero

Microbiology and Cytokine Levels Around Healthy Dental Implants and Teeth

BACKGROUND Elicitation of the relationship of periodontopathogens and pro-inflammatory cytokines to bone resorption and formation is significant to a growing body of research known as osteoimmunology. It is essential that clinically healthy peri-implant and periodontal sites are studied to contribute comparison data for investigations that are addressing diseased sites. PURPOSE The purpose of this study was to describe levels of selected pro-inflammatory cytokines in clinically healthy peri-implant and periodontal sites, and to examine whether cytokine levels may be related to specific bacterial/viral pathogens. MATERIALS AND METHODS Eleven subjects (mean age 56.2 +/- 10) participated in the study. Subgingival microbial samples were cultured for periodontopathic bacteria. Gingival crevicular fluid samples were analyzed by nested polymerase chain reaction for Cytomegalovirus (HCMV) and were tested for the quantification of Interleukin (IL)-8, IL-1beta, IL-6, IL-10, Tumor Necrosis Factor (TNF)-alpha, and IL-12p70 using flow cytometry (FACS). Findings for microbiota composition and cytokine levels were compared between implants and teeth (chi square, Kruskall-Wallis, Mann-Whitney; p < or = .05). RESULTS Both the frequency (%) and levels (%) of periodontopathic bacteria were higher around teeth than implants. The concentration (picogram per milliliter) of cytokines was more prominent around implants than teeth, reaching nearly twofold differences in some instances. Cytokine levels were higher when the sites analyzed were positive for any bacteria tested. HCMV was not detected. CONCLUSIONS Pro-inflammatory cytokine production was unrelated to heavy bacterial challenge. Nevertheless, when periodontopathic bacteria were detected by culture, cytokine levels were increased around both implants and teeth. Studies are needed to investigate the pro-inflammatory cytokines (especially IL-1beta and TNF-alpha) produced in spite of minimal bacterial accumulation.

Periodontal disease severity is related to high levels of C-reactive protein in pre-eclampsia.

Objective Recent studies have shown that pre-eclamptic women present a high prevalence of periodontitis, suggesting that active periodontal disease may play a role in the pathogenesis of pre-eclampsia. The present study analysed the effect of periodontal disease in the concentrations of serum high-sensitivity C-reactive protein (hs-CRP), and its association with pre-eclampsia. Methods A case–control study was carried out in Cali-Colombia, comprised of 398 pregnant women (145 cases and 253 controls) who were believed to have periodontal disease, between 28 and 36 weeks of gestational age. Pre-eclampsia cases were defined as blood pressure ≥ 140/90 mmHg and proteinuria ≥ 0.3 g/24 h. Controls were pregnant women with normal blood pressure, without proteinuria, matched by maternal age, gestational age and body mass index. Sociodemographic data, obstetric risk factors, periodontal state, subgingival microbial composition and hs-CRP levels were determined in both groups. Results The case and control groups were comparable for sociodemographic characteristics. In women with pre-eclampsia and confirmed periodontal disease (n = 138), hs-CRP levels increased according to the severity of the disease (gingivitis median 4.14 mg/dl; mild periodontitis median 4.70 mg/dl; moderate/severe periodontitis median 8.8 mg/dl; P = 0.01). A similar tendency was observed in controls with periodontal disease (n = 251), but it did not reach statistical significance (gingivitis median 5.10 mg/dl; mild periodontitis median 5.12 mg/dl; moderate/severe periodontitis median 6.90 mg/dl; P = 0.07). A significant difference in hs-CRP levels was observed in pre-eclamptic women with moderate/severe periodontitis compared to controls (P = 0.01). Conclusion These findings suggest that chronic periodontitis may increase hs-CRP levels in pregnant women and lead to complications such as pre-eclampsia.

Profiling of inflammatory cytokines produced by gingival fibroblasts after human cytomegalovirus infection

INTRODUCTION The purpose of this study was to determine the profile of inflammatory cytokines that are produced after in vitro infection of gingival fibroblasts with human cytomegalovirus (HCMV). MATERIALS AND METHODS Gingival fibroblasts were infected with the Towne strain of HCMV and the cytokine profile in the supernatant was studied using a human inflammation antibody array. Expression of messenger RNA (mRNA) using reverse transcription-polymerase chain reaction for interleukin-1beta (IL-1beta) and tumor necrosis factor-alpha (TNF-alpha) was also analyzed in infected gingival fibroblasts and gingival specimens from subjects with and without periodontitis according to HCMV detection. HCMV was determined in subgingival samples by nested polymerase chain reaction. RESULTS Gingival fibroblasts produced mainly IL-1alpha, IL-12p40, IL-12p70, IL-6, TNF-alpha, and IL-1beta after HCMV infection. Expression of mRNA for IL-1beta and TNF-alpha was increased after HCMV infection. Production of IL-1beta and TNF-alpha was increased in HCMV-positive periodontitis specimens. In addition, infected gingival fibroblasts produced more IL-8, monocyte chemoattractant protein 1, macrophage inflammatory proteins 1alpha, and 1beta over time postinfection in comparison to baseline. The lowest production of all cytokines studied corresponded to IL-2, IL-4, IL-13, and interferon-gamma. A decreasing production pattern was observed for granulocyte-macrophage colony-stimulating factor, IL-7, and IL-17 while IL-11 and macrophage colony-stimulating factor were increased at 72 h postinfection. CONCLUSIONS HCMV infection in gingival fibroblasts upregulated the production of proinflammatory-related cytokines and chemokines. The expression of IL-1beta and TNF-alpha was increased both in vitro and in specimens from HCMV-positive subjects with periodontitis. The overproduction of proinflammatory cytokines and chemokines as a result of viral infection should be considered an important pathogenic mechanism linking HCMV to periodontitis in vivo.

Comparison of nested polymerase chain reaction (PCR), real‐time PCR and viral culture for the detection of cytomegalovirus in subgingival samples

INTRODUCTION The purpose of this study was to compare nested polymerase chain reaction (PCR), real-time PCR, and shell vial for the detection of human cytomegalovirus (HCMV) in subgingival samples in periodontitis patients. METHODS A group of 44 patients and 24 individuals without periodontitis were included in the study. A full periodontal examination was conducted in each subject. Gingival crevicular fluid (GCF) was collected by pocket lavage and used for viral culture (shell vial). Additional subgingival samples were obtained with paper points and used for molecular analysis. Nested PCR and real-time PCR were used to detect and quantify HCMV. Students t-test and chi-squared test were used to compare groups. The sensitivity and specificity for the tests were calculated on 2 x 2 tables considering the nested PCR as the gold standard. RESULTS The detection of HCMV was greater using nested PCR than with either real-time PCR or shell vial (P < 0.0001). However, the frequency detection of both molecular techniques was higher than in viral culture (P < 0.0001). Only one case of chronic periodontitis was positive by viral culture. Agreement between nested PCR and real-time PCR was observed 47.7% and 4.1% of the time in the periodontitis and control groups, respectively. The sensitivity of real-time PCR was 60%, compared with 2.8% for the shell vial technique. CONCLUSIONS In conclusion, this study confirmed that active HCMV infection occurs in human periodontitis; however, its frequency seems to be low. In contrast, latent periodontal HCMV infection seems to be a more frequent event.

Biology and pathogenesis of cytomegalovirus in periodontal disease

Abstract Human periodontitis is associated with a wide range of bacteria and viruses and with complex innate and adaptive immune responses. Porphyromonas gingivalis, Tannerella forsythia, Aggregatibacter actinomycetemcomitans, Treponema denticola, cytomegalovirus and other herpesviruses are major suspected pathogens of periodontitis, and a combined herpesvirus–bacterial periodontal infection can potentially explain major clinical features of the disease. Cytomegalovirus infects periodontal macrophages and T‐cells and elicits a release of interleukin‐1β and tumor necrosis factor‐α. These proinflammatory cytokines play an important role in the host defense against the virus, but they also have the potential to induce alveolar bone resorption and loss of periodontal ligament. Gingival fibroblasts infected with cytomegalovirus also exhibit diminished collagen production and release of an increased level of matrix metalloproteinases. This article reviews innate and adaptive immunity to cytomegalovirus and suggests that immune responses towards cytomegalovirus can play roles in controlling, as well as in exacerbating, destructive periodontal disease.

The Profile of Inflammatory Cytokines in Gingival Crevicular Fluid around Healthy Osseointegrated Implants

OBJECTIVE Regardless of gingival health and subgingival microbiology, production of cytokines within peri-implant tissues may be different from that of teeth. The objective of this study was to describe the peri-implant levels of pro-inflammatory cytokines and subgingival microbiology in clinically healthy sites. MATERIALS AND METHODS Subgingival plaque and gingival crevicular fluid (GCF) were obtained from 28 clinically healthy implants and 26 teeth selected from 24 individuals. Microbial composition was determined by selective anaerobic culture techniques. Pro-inflammatory cytokines were quantified by flow cytometry analysis of GCF. The concentration of cytokines between implants and teeth were compared with the independent t-test. RESULTS The concentration of cytokines was higher in GCF from healthy implants than in teeth. The profile of cytokines was characteristic of an innate immune response. A more frequent detection of periodontopathic bacteria was observed in teeth than implants. Cultivable levels of periodontopathic bacteria were similar between implants and teeth. CONCLUSIONS Despite gingival tissue health and scarce plaque accumulation, the profile of inflammatory cytokines in implant crevicular fluid was distinctive of an innate immune response and in higher concentration than in teeth. Other than bacterial stimulus, intrinsic factors related to implants may account for more cytokine production than teeth.

Periodontal disease in children and adolescents of Latin America

Periodontal diseases are a group of infectious diseases that mainly include gingivitis and periodontitis. Gingivitis is the most prevalent form of periodontal disease in subjects of all ages, including children and adolescents. Less frequent types of periodontal disease include aggressive periodontitis, acute necrotizing ulcerative gingivitis and various diseases of herpesviral and fungal origin. This review aimed to retrieve relevant information from Latin America on the prevalence of periodontal diseases among children and adolescents of the region. Gingivitis was detected in 35% of young Latin American subjects and showed the highest frequencies in Colombia (77%) and Bolivia (73%) and the lowest frequency in Mexico (23%). The frequency of gingivitis in subjects from other Latin American countries was between 31% and 56%. Periodontitis may affect <10% of the young population in Latin America, but the data are based on only a few studies. A more precise assessment of the distribution and severity of periodontal disease in children and adolescents of Latin America may help policy makers and dentists to institute more effective public health measures to prevent and treat the disease at an early age to avoid major damage to the permanent dentition.

Effects of cytomegalovirus infection on the mRNA expression of collagens and matrix metalloproteinases in gingival fibroblasts

BACKGROUND AND OBJECTIVE The purpose of this in vitro investigation was to study the effects of human cytomegalovirus infection on the mRNA expression for collagens I and III and for matrix metalloproteinases 1 and 2 in gingival fibroblasts. MATERIAL AND METHODS Gingival fibroblasts were experimentally infected with the Towne strain of human cytomegalovirus and the kinetics of expression of mRNA for collagens I and III and for matrix metalloproteinases 1 and 2 was studied at different time-points. Total RNA was isolated at the indicated time, and the reverse transcription-polymerase chain reaction was used to analyze the level of mRNA expression. In addition, gingival specimens were obtained from 14 periodontitis and from three non-periodontitis subjects and mRNA analysis for collagens and metalloproteinases was carried out. Nested polymerase chain reaction was used to determine the presence or absence of human cytomegalovirus in subgingival samples from each subject. RESULTS The infection of gingival fibroblasts with human cytomegalovirus during a 0-72-h period resulted in progressive reduction in the expression of mRNA for collagens I and III (p < 0.05). A higher concentration of human cytomegalovirus resulted in varying degrees of mRNA reduction, suggesting a virally mediated effect. Biopsies from human cytomegalovirus-positive individuals with periodontitis had a higher expression of mRNA for collagens I and III than biopsies from human cytomegalovirus-negative individuals. An up-regulation in the mRNA expression of matrix metalloproteinases 1 and 2 over time was observed (p < 0.05). Analysis of mRNA expression in gingival biopsies demonstrated higher expression of matrix metalloproteinase-1 in human cytomegalovirus-positive periodontitis specimens compared with human cytomegalovirus-negative periodontitis specimens. CONCLUSION Altered expression of mRNA for collagens and metalloproteinases in human cytomegalovirus-infected gingival fibroblasts should be considered as possible modifying mechanisms in periodontitis-infected sites.

Determinantes del Diagnóstico Periodontal

Resumen Un correcto diagnostico periodontal es necesario para la realizacion de una terapia periodontal exitosa en nuestro paciente. Entendiendo que la enfermedad periodontal es un proceso infeccioso-inflamatorio, diferentes variables se deben analizar clinicamente para determinar el diagnostico. El diagnostico entonces es un analisis concienzudo de la expresion clinica de la enfermedad, desde gingivitis hasta periodontitis. Este articulo analiza los determinantes mas importantes del diagnostico periodontal utilizados en la practica clinica diaria.

Periodontal intervention effects on pregnancy outcomes in women with preeclampsia

Objective: To determine the efficacy of periodontal intervention on pregnancy outcome in mild preeclamptic women. Methods: A sample of 60 pregnant women with mild preeclampsia (blood pressure levels <160/110 mm and proteinuria >300 mg/l in 24 hours urine) from the Hospital Universitario del Valle (Cali, Colombia) was included to the study. Preeclamptic women were randomized in two groups, one with periodontal intervention (PIG, N=28) and another in which the periodontal intervention was practiced after childbirth (NPIG, N=32). Maternal socio-demographic, medical and periodontal data were obtained. PIG included patients in which supragingival and subgingival cleaning within ultrasonic and manual devices were performed after study inclusion. The progression from mild to severe preeclampsia, eclampsia or HELLP syndrome, the number of days of clinical stability and the percentile of birth-weight adjusted for gestational age were evaluated in both groups. Results: Most of the patients (60%) were multigravids. Gestational age at inclusion was 31.8±1.6 weeks. Chronic periodontitis was a frequent finding (61.7%). Social, demographic, medical and periodontal conditions were similar between both groups. Disease progression to severe preeclampsia, eclampsia or HELLP syndrome was also similar (89.2% PIG versus 84.4%, p=0.65) (OR=1.06 IC 95% 0.87-1.29, p=0.65). Days of clinical stability were similar between the groups (median 10 days , range 1-46, PIG versus 12 days, range 1-59, p=0.57) and the percentile of birth weight adjusted with gestational age had no differences between the groups (median percentil 50 range 5-90 PIG versus percentil 55 range 5-95, p=0.73). Conclusion: Periodontal intervention does not seem to harm the health, the severity or alter the frequency on maternal complications in mild preeclampsia subjects.