Overexpression of proinflammatory cytokines in dental pulp tissue and distinct bacterial microbiota in carious teeth of Mexican Individuals.

The prevalence of dental caries in the Mexican adult population aged 20 to 85 years is around 93.3%, and 50% in Mexican children and adolescents. Worldwide, it is the most common non-communicable disease. One of the main etiological factors for dental caries is the oral microbiome and changes in its structure and function, with an expansion of pathogenic bacteria like Streptococcus mutans. The exposed dental pulp tissue triggers an innate immune response to counteract this bacterial invasion. The relation between oral dysbiosis and innate immune responses remains unclear. We aimed to understand the relationship between innate immune response and the oral microbiota by quantifying the expression of Toll-like receptors (TLRs) and proinflammatory markers (cytokines and a chemokine) in dental pulp tissue, either exposed or not to carious dentin, and to correlate this information with the oral microbiome found in healthy teeth and those with moderate caries. RNA was purified from pulp tissue, subjected to RT-qPCR and analysed with the ΔΔCt method. Supragingival dental plaque of non-carious teeth and dentin of carious teeth were subjected to 16S targeted sequencing. Principal coordinate analysis, permutational multivariate ANOVA, and linear discriminant analysis were used to assess differences between non-carious and carious teeth. Correlations were assessed with Spearman´s test and corrected for multiple comparisons using the FDR method. The relative abundance (RA) of Lactobacillus, Actinomyces, Prevotella, and Mitsuokella was increased in carious teeth; while the RA of Haemophilus and Porphyromonas decreased. Olsenella and Parascardovia were only detected in carious teeth. Significant overexpression of interleukin 1 beta (IL1 ß), IL6, and CXCL8 was detected in pulp tissue exposed to carious dentin. IL1ß correlated positively with TLR2 and Actinomyces; yet negatively with Porphyromonas. These findings suggest that immune response of pulp tissue chronically exposed to cariogenic microbiome is triggered by proinflammatory cytokines IL1ß and IL6 and the chemokine CXCL8.

Antimicrobial Photodynamic Therapy Associated with Conventional Endodontic Treatment: A Clinical and Molecular Microbiological Study.

This study evaluated antimicrobial photodynamic therapy (aPDT) as an adjunct to endodontic treatment. Ten uniradicular teeth (control group (CG) = 4 (2 and test group (TG) = 6) with primary endodontic infections, from both genders, between 17 and 65 years old, were analyzed. Microbiological samples were collected before and after chemical-mechanical instrumentation (CMI), after aPDT (for the TG), and after the removal of the temporary restorations (second session). In TG, the aPDT was performed with 100 µg mL-1 methylene blue and irradiated with low power laser (InGaAIP, 660 nm; 100 mW; 40 s) with a fiber-coupled optical laser. Another irradiation (3 J; 30 s; spot size of 3 mm2 ) was performed in the gingiva close to the apical foramen. The PCR was performed, after previous whole-genome amplification, for Enterococcus faecalis, Candida genus and Bacteria domain. For TG, a positive tooth for Candida spp. before of the CMI presented negative results in subsequent samples. Additionally, E. faecalis species was present in four samples before CMI, two after CMI, in one after the aPDT and was not detected at the second session. aPDT may be an effective adjunct therapy, resulting in a reduction (P = 0.0286) of the incidence of E. faecalis before root canal obturation.

In vitro evaluation of the antimicrobial effects of different intracanal medications in necrotic immature teeth.

AIM: This was to assess the antimicrobial effectiveness of pastes used as intracanal medicaments against Enterococcus faecalis in necrotic immature teeth. METHOD: An agar-disc diffusion method over a period of 30 days was used. The following medications were analysed: triple antibiotic paste (TAP) I; double antibiotic paste (DAP) I; DAP I with calcium hydroxide; calcium hydroxide paste with saline solution 0.9 %; calcium hydroxide paste with chlorhexidine 2 % gel; DAP II; DAP II with zinc oxide; DAP II with calcium hydroxide; and saline solution 0.9 % as control. The diameters of the halos of inhibition (in mm) of tested medicaments were determined and analysed with one-way analysis of variance and Tukey's post hoc tests. RESULTS: TAP I (G1) was significantly more effective against E. faecalis, and showed the largest halo of inhibition during all the experiments. DAP groups (G2 and G6) also showed inhibiting bacterial growth with said inhibition remaining stable throughout the 30-day period. In contrast, G4 and G5 groups, in which Ca(OH)2 was added to saline and chlorhexidine, had no antibacterial effect. CONCLUSIONS: TAP and DAP showed better antibacterial efficacy and remained active for 30 days. Combination of Ca(OH)2 with antibiotics should be avoided, due to the possibility of antibiotic hydrolysis.

Immediate human pulp response to ethanol-wet bonding technique.

OBJECTIVES: To evaluate the short-term response of human pulps to ethanol-wet bonding technique. METHODS: Deep class V cavities were prepared on 17 sound premolars and divided into three groups. After acid-etching, the cavities from groups 1 (G1) and 2 (G2) were filled with 100% ethanol or distilled water, respectively, for 60 s before the application of Single Bond 2. In group 3 (G3, control), the cavity floor was lined with calcium hydroxide before etching and bonding. All cavities were restored with resin composite. Two teeth were used as intact control. The teeth were extracted 48h after the clinical procedures. From each tooth serial sections were obtained and stained with haematoxylin and eosin (H/E) and Masson's trichrome. Bacteria microleakage was assessed using Brown & Brenn. All sections were blindly evaluated for five histological features. RESULTS: Mean remaining dentine thickness was 463±65µm (G1); 425±184µm (G2); and 348±194µm (G3). Similar pulp reactions followed ethanol- or water-wet bonding techniques. Slight inflammatory responses and disruption of the odontoblast layer related to the cavity floor were seen in all groups. Stained bacteria were not detected in any cavities. Normal pulp tissue was observed in G3 except for one case. CONCLUSIONS: After 48h, ethanol-wet bonding does not increase pulpal damage compared to water-wet bonding technique. CLINICAL SIGNIFICANCE: Ethanol-wet bonding may increase resin-dentine bond durability. This study reported the in vivo response of human pulp tissue when 100% ethanol was applied previously to an etch-and-rinse simplified adhesive system.

Microbial reduction by two chemical-mechanical protocols in primary teeth with pulp necrosis and periradicular lesion - an in vivo study.

The objective of this study was to determine the efficacy of chemical-mechanical procedures of two endodontic protocols for septic content reduction of root canals from primary teeth with pulp necrosis and periradicular lesion. Twenty-four primary root canals with pulp necrosis and periradicular lesion were divided into two treatment groups (n=12): multiple-visit and single-visit protocols. Samples were collected using sterile paper points before and after endodontic cleaning followed by microbiological identification through checkerboard DNA-DNA hybridization. Statistical analysis was performed using Proportion Test for score=0 comparing the findings before and after treatment for each group (Wilcoxon's test) as well as the differences in scores between protocols (Mann-Whitney's test) (p<0.05). Data were expressed as prevalence (presence or absence) and estimate of the average count (x10(5) cells) of each species. Differences in proportions of score=0 prior to treatment were non-significant (p=0.415), demonstrating equivalence between groups. A significant increase in score=0 was detected after treatment for both groups (p<0.0001). Single-visit protocol achieved a significantly greater reduction in mean scoring following endodontic treatment (p=0.024). Both protocols were capable of significantly reducing septic content in root canals of primary teeth with periradicular lesion. Moreover, single-visit protocol showed greater efficacy in reducing endodontic infection.

Microbial Reduction by Two Chemical-Mechanical Protocols in Primary Teeth with Pulp Necrosis and Periradicular Lesion - An In Vivo Study

The objective of this study was to determine the efficacy of chemical-mechanical procedures of two endodontic protocols for septic content reduction of root canals from primary teeth with pulp necrosis and periradicular lesion. Twenty-four primary root canals with pulp necrosis and periradicular lesion were divided into two treatment groups (n=12): multiple-visit and single-visit protocols. Samples were collected using sterile paper points before and after endodontic cleaning followed by microbiological identification through checkerboard DNA-DNA hybridization. Statistical analysis was performed using Proportion Test for score=0 comparing the findings before and after treatment for each group (Wilcoxon's test) as well as the differences in scores between protocols (Mann-Whitney's test) (p<0.05). Data were expressed as prevalence (presence or absence) and estimate of the average count (x105 cells) of each species. Differences in proportions of score=0 prior to treatment were non-significant (p=0.415), demonstrating equivalence between groups. A significant increase in score=0 was detected after treatment for both groups (p<0.0001). Single-visit protocol achieved a significantly greater reduction in mean scoring following endodontic treatment (p=0.024). Both protocols were capable of significantly reducing septic content in root canals of primary teeth with periradicular lesion. Moreover, single-visit protocol showed greater efficacy in reducing endodontic infection.

O objetivo deste estudo foi determinar a eficácia das manobras químico-mecânicas de dois protocolos endodônticos, na redução do conteúdo séptico de canais radiculares de dentes decíduos com polpa necrosada e lesão perirradicular. Vinte e quatro canais radiculares decíduos com necrose pulpar e lesão perirradicular foram divididos em dois grupos de tratamento (n=12): multisessões e sessão única. Amostras foram coletadas usando pontas de papel estéreis, antes e após a limpeza endodôntica, seguido de identificação microbiológica por hibridização DNA-DNA checkerboard. A análise estatística foi realizada usando teste de proporções para escore=0, comparando os achados antes e após tratamento para cada grupo (teste de Wilcoxon) e as diferenças dos escores entre os protocolos (teste de Mann-Whitney) (p<0,05). Os dados foram expressos em prevalência (presença ou ausência) e contagem média (x105 células) de cada espécie. As diferenças nas proporções de escore=0 antes do tratamento não foram significativas (p=0,415), mostrando equivalência entre os grupos. Um aumento significativo de escore=0 foi detectado após o tratamento para ambos os grupos (p<0,0001). O protocolo de sessão única mostrou uma redução significativamente maior dos escores médios após o tratamento endodôntico (p=0,024). Ambos os protocolos são capazes de reduzir significativamente o conteúdo séptico de canais radiculares de dentes decíduos com lesão perirradicular. Entretanto, o protocolo de sessão única mostrou uma maior eficácia na redução da infecção endodôntica.

Clinical antimicrobial efficacy of NiTi rotary instrumentation with NaOCl irrigation, final rinse with chlorhexidine and interappointment medication: a molecular study.

AIM: To evaluate clinically the antibacterial effects of root canal treatment procedures using molecular microbiology analyses. METHODOLOGY: Samples were taken from 14 necrotic root canals of teeth with apical periodontitis before (S1) and after instrumentation with NaOCl irrigation (S2), a final rinse with chlorhexidine (CHX) (S3) and then one-week interappointment medication with calcium hydroxide/CHX paste (S4). The parameters examined included the following: incidence of positive broad-range PCR results for bacterial presence; impact on bacterial community structures evaluated by PCR-Denaturing Gradient Gel Electrophoresis (DGGE); quantitative bacterial reduction determined by real-time PCR; and identification of bacterial persisters by cloning and sequencing. Data from the different tests were subjected to statistical analyses and diversity indicator calculations. RESULTS: All S1 samples were positive for bacteria in all tests. Treatment procedures promoted a decrease in microbial diversity and significantly reduced the incidence of positive results and the bacterial counts (P < 0.05). In general, each subsequent treatment step improved disinfection. No specific taxon or community pattern was associated with post-treatment samples. CONCLUSION: Supplementary steps consisting of a final rinse with CHX followed by calcium hydroxide interappointment medication promoted further decrease in the bacterial bioburden to levels significantly below those achieved by the chemomechanical procedures alone. Because the long-term outcome of root canal treatment is dependent upon maximal bacterial reduction, the present results are of clinical relevance.

Partial caries removal in primary teeth: association of clinical parameters with microbiological status.

The relationship between clinical characteristics of carious dentin and bacterial colonization after partial caries removal is not completely understood. The aim of this study was to compare microbial counts between categories of carious dentin color, consistency and humidity, and to evaluate the correlation between these characteristics and the presence of cariogenic microorganisms in deep cavities (2/3 or more of the dentin thickness) submitted to partial caries removal. Sixteen primary teeth were submitted to the removal of all carious tissue from the lateral walls of the cavity, whereas carious tissue of the pulp wall was removed superficially. Dentin in the pulp wall was classified according to color, consistency and humidity immediately after cavity preparation and 3-6 months after cavity sealing and a tissue sample was collected on the same occasion for microbiological evaluation. Before sealing, Streptococcus mutans (p = 0.033) and Lactobacillus spp. (p = 0.048) counts were higher in cavities with humid dentin compared to cavities with dry dentin. A negative correlation was observed between carious dentin consistency and S. mutans count during this phase (r(s) = -0.571; p = 0.020). Arrest of dentinal caries lesions was observed after sealing, which was characterized by a reduction of bacterial counts and changes in dentin color, consistency and humidity, irrespectively of baseline dentin characteristics. The clinical characteristics of carious dentin change after the period of cavity sealing and cannot be applied as absolute indicators to limit the excavation of carious dentin when minimally invasive techniques are used.

Response of human dental pulp to calcium hydroxide paste preceded by a corticosteroid/ antibiotic dressing agent

Aim: To evaluate the treatment with corticosteroid/antibiotic dressing in pulpotomy with calciumhydroxide. Methods: Forty-six premolars were pulpotomized and randomly assigned into 3groups. In Group I pulpal wound was directly capped with calcium hydroxide, and Group II and Group III received corticosteroid/antibiotic dressing for 10 min or 48 h, respectively, before pulp capping. Teeth were processed for histological analysis after 7, 30 or 60 days to determine inflammatory cell response, tissue disorganization, dentin bridge formation and presence of bacteria.Attributed scores were analyzed by Kruskal-Wallis and Mann-Whitney tests (á=0.05). Results:On the 7th day, all groups exhibited dilated and congested blood vessels in the tissue adjacent to pulpal wound. The inflammatory cell response was significantly greater in Group III (p<0.05). On the 30th day, in all groups, a thin dentin matrix layer was deposited adjacent to the pulpal wound and a continuous odontoblast-like cell layer underlying the dentin matrix was observed. On the60th day, all groups presented a thick hard barrier characterized by an outer zone of dystrophic calcification and an inner zone of tubular dentin matrix underlined by a defined odontoblast-like celllayer. Conclusions: Within the limitations of present study, considering that the treatment was performed in healthy teeth, it may be concluded that the use of a corticosteroid/antibiotic dressing before remaining tissue protection with calcium hydroxide had no influence on pulp tissue healing.

Heat-killed Enterococcus faecalis alters nitric oxide and CXCL12 production but not CXCL8 and CCL3 production by cultured human dental pulp fibroblasts.

INTRODUCTION: Fibroblasts are the most abundant cells in dental pulp. To investigate their capacity to produce the chemokines CCL3, CXCL8, and CXCL12 as well as nitric oxide (NO), we evaluated the production of these mediators in supernatants of cultured human dental pulp fibroblasts (HDPF) stimulated by heat-killed Enterococcus faecalis (HKEF). METHODS: Primary cultures of HDPF were stimulated with medium alone or HKEF (1:1, 10:1, or 100:1 bacteria:fibroblast) for 1, 6, and 24 hours. Chemokines and NO were assessed through enzyme-linked immunosorbent assay and Griess reaction, respectively. Statistical analysis was performed by using analysis of variance and Tukey post test. RESULTS: CCL3 was not detected, whereas constitutive CXCL8 was not affected. Production of CXCL12 was increased at 1 and 6 hours, and NO was increased at the concentration of 1:1 bacteria:fibroblast at 24 hours. Viability and proliferation assays did not reveal cell number differences. CONCLUSIONS: These findings demonstrate that heat-killed E. faecalis is able to increase production of CXCL12 and NO by HDPF.

Microbiological and microscopic analysis of the pulp of non-vital traumatized teeth with intact crowns.

OBJECTIVE: This study evaluated the presence of microorganisms and analyzed microscopically the pulp of 20 traumatized human teeth with intact crowns and clinical diagnosis of pulp necrosis, based on the association of at least three of the clinical criteria: crown discoloration, negative response to thermal and electric pulp vitality tests, positive response to vertical and horizontal percussion, pain on palpation or mobility. MATERIAL AND METHODS: Microbiological collection was performed from the root canals to evaluate the presence of microorganisms. The pulp samples were stained with hematoxylin and eosin (H.E.) for histological evaluation of possible morphological alterations. RESULTS: Analysis of results was performed by statistical tests (linear regression test and diagnostic analysis) and subjective analysis of the sections stained with H.E. and revealed that only 15% of the sample did not exhibit microbial development. The time elapsed between dental trauma and onset of endodontic intervention ranged from 15 days to 31 months; the percussion test presented high sensitivity (80%) for detection of microorganisms in the root canal of traumatized teeth; 3 teeth (15%) did not present pulp tissue, being characterized as complete autolysis; analysis of pulp samples was performed on the other 17 cases, among which 3 (15%) exhibited partial necrosis without possibility of repair and 14 presented complete necrosis; none of the clinical criteria employed for the diagnosis of pulp necrosis in traumatized teeth was pathognomonic. CONCLUSIONS: The present results allowed the following conclusions: with regard to microbiological findings, 85% of teeth presented microorganisms in the root canal, despite the presence of an intact crown. Concerning the microscopic findings, 100% of traumatized teeth presented pulp necrosis; the pulp vitality tests based on pulp response to heat, cold and vertical percussion were the most reliable to diagnose pulp necrosis in traumatized teeth.

Microbiological and microscopic analysis of the pulp of non-vital traumatized teeth with intact crowns

OBJECTIVE: This study evaluated the presence of microorganisms and analyzed microscopically the pulp of 20 traumatized human teeth with intact crowns and clinical diagnosis of pulp necrosis, based on the association of at least three of the clinical criteria: crown discoloration, negative response to thermal and electric pulp vitality tests, positive response to vertical and horizontal percussion, pain on palpation or mobility. MATERIAL AND METHODS: Microbiological collection was performed from the root canals to evaluate the presence of microorganisms. The pulp samples were stained with hematoxylin and eosin (H.E.) for histological evaluation of possible morphological alterations. RESULTS: Analysis of results was performed by statistical tests (linear regression test and diagnostic analysis) and subjective analysis of the sections stained with H.E. and revealed that only 15 percent of the sample did not exhibit microbial development. The time elapsed between dental trauma and onset of endodontic intervention ranged from 15 days to 31 months; the percussion test presented high sensitivity (80 percent) for detection of microorganisms in the root canal of traumatized teeth; 3 teeth (15 percent) did not present pulp tissue, being characterized as complete autolysis; analysis of pulp samples was performed on the other 17 cases, among which 3 (15 percent) exhibited partial necrosis without possibility of repair and 14 presented complete necrosis; none of the clinical criteria employed for the diagnosis of pulp necrosis in traumatized teeth was pathognomonic. CONCLUSIONS: The present results allowed the following conclusions: with regard to microbiological findings, 85 percent of teeth presented microorganisms in the root canal, despite the presence of an intact crown. Concerning the microscopic findings, 100 percent of traumatized teeth presented pulp necrosis; the pulp vitality tests based on pulp response to heat, cold and vertical percussion ...

Evaluation of two mineral trioxide aggregate compounds as pulp-capping agents in human teeth.

AIM: The present randomized, controlled prospective study evaluated the histomorphological response of human dental pulps capped with two grey mineral trioxide aggregate (MTA) compounds. METHODOLOGY: Pulp exposures were performed on the occlusal floor of 40 human permanent pre-molars. The pulp was capped either with ProRoot (Dentsply) or MTA-Angelus (Angelus) and restored with zinc oxide eugenol cement. After 30 and 60 days, teeth were extracted and processed for histological examination and the effects on the pulp were scored. The data were subjected to Kruskal-Wallis and Conover tests (alpha = 0.05). RESULTS: In five out of the 40 teeth bacteria were present in pulp tissue. No significant difference was observed between the two materials (P > 0.05) in terms of overall histological features (hard tissue bridge, inflammatory response, giant cells and particles of capping materials). Overall, 94% and 88% of the specimens capped with MTA-Angelus and ProRoot, respectively, showed either total or partial hard tissue bridge formation (P > 0.05). CONCLUSIONS: Both commercial materials ProRoot (Dentsply) and MTA-Angelus (Angelus) produced similar responses in the pulp when used for pulp capping in intact, caries-free teeth.

Occurrence of herpes simplex virus 1 and three periodontal bacteria in patients with chronic periodontitis and necrotic pulp.

Viral and bacterial associations appear to be implicated in the development of periodontal infections. Little information is available describing the periodontopathic agents in root canals with necrotic pulp. In this study, the occurrence and the combinations among herpes simplex virus type 1 (HSV-1) and Dialister pneumosintes, Tannerella forsythia, and Treponema denticola in patients with chronic periodontitis and necrotic pulp were evaluated. Clinical samples from healthy subjects and patients with periodontal or pulp infections were analyzed using a nested polymerase chain reaction PCR to detect HSV and PCR to detect the 3 periodontal bacteria. The presence of Tannerella forsythia and Treponema denticola was observed in healthy, periodontitis, and necrotic pulp patients. HSV was observed in periodontitis and necrotic pulp patients, and no healthy subject harbored D. pneumosintes or HSV. The occurrence of Tannerella forsythia was not statistically significant in patients with necrotic pulp (P = 0.704). Periodontal bacteria were observed varying from 10.3% to 20.7% in periodontitis and necrotic pulp patients. The presence of Treponema denticola - HSV association was predominant in patients showing necrotic pulp (24.1%); however, HSV alone was observed in one patient with periodontitis and in another patient with necrotic pulp. The presence of double association among bacteria or bacteria - HSV could indicate a role in both periodontitis and necrotic pulp, and Tannerella forsythia - Treponema denticola - HSV and Tannerella forsythia - D. pneumosintes - Treponema denticola - HSV associations might be important in periodontitis.

Histopathologic study of physiological and pathological resorptions in human primary teeth.

This study presents a histological analysis through optical microscopy of primary teeth with physiological and pathological resorptions to outline the histological profile of resorptions. Sixty teeth were examined: 19 primary teeth with physiological resorption and 41 primary teeth with pathological resorption. To analyze the histological conditions of the pulp, periradicular tissue, and the resorption areas, and to investigate the presence, intensity, and location of bacteria, slides were prepared using the hematoxylin-eosin and the Brown-Brenn techniques. For the teeth with physiological resorption, normal pulps and no evidence of bacteria were found. For the teeth with pathological resorption, pulpal alterations, atypical resorption, and bacteria were observed.

Bacteria recovered from dental pulp induce apoptosis of lymph node cells.

Apoptosis is critical in the pathogenesis of several infectious diseases. The induction of apoptosis was assessed in mouse lymph node cells by four bacteria recovered from infected human dental pulp: Gemella morbillorum, Clostridium butyricum, Fusobacterium nucleatum and Bifidobacterium adolescentis. Smaller lymph nodes and smaller numbers of cells were observed after experimental dental pulp infection with C. butyricum, suggesting that this bacterium induces cell death. Apoptosis was evaluated by determination of cell ploidy and detection of DNA degradation in cells cultured with killed bacteria. Paraformaldehyde-killed C. butyricum and heat-killed G. morbillorum induced substantial cell death, while F. nucleatum and B. adolescentis induced cell death at lower levels. No bacterial preparations induced apoptosis in cells from mice genetically deficient for tumour necrosis factor receptor p55 (TNFRp55), implicating this receptor directly or indirectly as a mediator in the process. It was concluded that apoptosis may be induced during periapical lesions of pulpal origin.

The reaction of dental pulp to Escherichia coli lipopolysaccharide and Enterococcus faecalis lipoteichoic acid

This research evaluates the effects of the lipopolysaccharides (LPS) from Escherichia coli and lipoteichoic acid (LTA) from Enterococcus faecalis on dental pulp. These molecules are components of the Gram-negative and Gram-positive bacteria cell wall, respectively. Ten dogs were used in the experiment. Inoculation in surgically opened pulp and coronal restoration with glass ionomer was the method chosen. The evaluation times were 1, 7, 15, 30 and 60 days. The results showed that the LPS and LTA, at 150 icg/ml, produced a negative interference in the pulp leading to destruction. LTA caused less damage than LPS.

Human pulpal response to direct pulp capping with an adhesive system.

PURPOSE: To evaluate clinically and microscopically the human pulp response when directly capped with an adhesive system or calcium hydroxide over short (9-12 days) and long (53-204 days) experimental periods. MATERIALS AND METHODS: Fifty-one sound human premolars scheduled for orthodontic extraction, had their pulp horns gently exposed with a diamond point. Debris in the pulp wound was washed out with a sterile saline solution. The pulps were then capped with either an adhesive system (Scotchbond Multi-Purpose Plus) or calcium hydroxide. All teeth were subsequently restored with resin-based composite (Z-100) according to the manufacturer's instructions. After the experimental periods, the teeth were extracted and processed for light microscopic examination. RESULTS: Short-term: the pulp tissue capped with SBMP-P exhibited dilated and congested blood vessels associated with a moderate inflammatory response and blanching of pulp cell nuclei. Long-term: no evidence of healing and bridge formation was observed. A persistent mild inflammatory pulp response was present. Micro-abscesses were detected in three cases associated with bacterial infiltration. Calcium hydroxide stimulated early pulp repair and dentin bridging which extended into the longest period.