Successful pulp revascularization of an autotransplantated mature premolar with fragile fracture apicoectomy and plasma rich in growth factors: a 3-year follow-up.

AIM: This case report demonstrates a positive outcome of the adjuvant use of fragile fracture (FF), which is a technique used to harvest dental pulp stem cells (DPSCs), and platelet-rich plasma (PRP) in a mandibular premolar (tooth 44) with a completely formed root that was transplanted into a surgically created socket and which maintained pulp vitality and function. SUMMARY: After virtual surgical planning, a 3D tooth replica of tooth 44 was fabricated. A surgical socket was created in the position of tooth 14; then, tooth 44 was extracted and the root dentine was abraded using a turbine diamond bur 3 mm from the apex until a circular groove was prepared around the outer circumference of the root; and then, an FF was performed without damaging the pulp tissue. PRP was placed in the socket, after which the donor tooth was inserted in the recipient area. At 2 weeks post-treatment, orthodontic traction was applied. At 3-year follow-up, the tooth had adequate alignment and was asymptomatic. Response to pulp testing was positive, and the presence of pulp canal obliteration was observed as a sign of pulpal healing. KEY LEARNING POINTS: Autotransplantation is a good alternative for replacing missing teeth, with repair of tissues and pulp revascularization. Revascularization of an autotransplanted mature tooth using the fragile fracture technique and PRP scaffold is a feasible option and might have positive effects on the long-term outcome of the procedure. Including completely formed teeth as donors in autotransplantation, maintaining vitality and their functions is an option that warrants further study.

Gene expression of growth factors with angiogenic potential in human dental pulp tissue from teeth with complete and incomplete root development.

AIM: To quantify the expression of angiogenic growth factors (ANG2, VEGFA, TGFß1) and their corresponding receptors (VEGFR1, VGFR2, NRP1 and TGFßR1) in human dental pulps from extracted third molars with complete and incomplete root development. METHODOLOGY: Fifty-six dental pulp samples obtained from freshly extracted human third molars were divided equally into two groups according to their stage of root development; 28 third molars with complete root development and 28 third molars with incomplete root development. All samples were processed and total RNA was extracted, cDNA was then synthetized for each sample and the target genes expression profiles for ANG2, VEGFA, VEGFR1, VEGFR2, NRP1, TGFß1 and TGFßR1 were obtained by RT2-PCR. The data was analysed with a Student's t-test to compare the replicate ∆∆Ct values for each gene. RESULTS: Teeth with incomplete root development were associated with a significantly greater gene expression of TGFßR1 (P = 0.03), whereas in teeth with complete root development the genes that had significantly greater expression were VEGFA (P = 0.04). CONCLUSION: The angiogenic growth factors (ANG2, VEGFA, TGFß1) and their receptors (NRP1, VEGFR1, VEGFR2 and TGFßR1) were expressed in pulps of teeth with complete and incomplete root development measured by RT2-PCR, with TGFBR1 genes being significantly different in teeth with incomplete root development and VEGFA genes in teeth with complete root development.

Substance P and Calcitonin gene-related peptide expression in human periodontal ligament after root canal preparation with Reciproc Blue, WaveOne Gold, XP EndoShaper and hand files.

AIM: To quantify Substance P (SP) and Calcitonin gene-related peptide (CGRP) expression in healthy human periodontal ligament from premolars after root canal preparation with Reciproc Blue, WaveOne Gold, XP EndoShaper and hand files. METHODOLOGY: A total of 50 human periodontal ligament samples were obtained from healthy mandibular premolars where extraction was indicated for orthodontic reasons. Prior to extraction, 40 of these premolars were equally divided into four groups, and root canals were prepared using four different systems: Reciproc Blue, WaveOne Gold, XP EndoShaper and a hand instrumentation technique. The remaining 10 healthy premolars were extracted without treatment and served as a negative control group. All periodontal ligament samples were processed, and SP and CGRP were measured by radioimmunoassay. The Kruskal-Wallis test was used to establish significant differences between groups and LSD post hoc comparisons were also performed. RESULTS: Greater SP and CGRP values were found in the hand instrumentation group, followed by the XP EndoShaper, WaveOne Gold and the Reciproc groups. The lower SP and CGRP values were for the healthy periodontal ligament group. The Kruskal-Wallis test revealed significant differences between groups (P < 0.05). Post hoc Least Significant Difference (LSD) tests revealed significant differences (P < 0.05) in SP and CGRP expression between all the comparisons except for the Reciproc Blue and WaveOne Gold group (P > 0.05). CONCLUSION: All the root canal preparation techniques tested increased SP and CGRP expression in human periodontal ligament, with hand files and XP EndoShaper instruments being associated with greater neuropeptide release compared to Reciproc Blue and WaveOne Gold files.

Angiogenic mechanisms of human dental pulp and their relationship with substance P expression in response to occlusal trauma.

Angiogenesis is the formation of new blood vessels based on a pre-existing vasculature. It comprises two processes, sprouting of endothelial cells and the division of vessels due to abnormal growth of the microvasculature. It has been demonstrated that substance P (SP) can induce angiogenesis either by modulating endothelial cell growth (direct mechanism) or by attracting cells with angiogenic potential to the injury site (indirect mechanism). Therefore, the purpose of this article is to review the angiogenic mechanisms that regulate mineralized tissue formation in human dental pulp tissue and their relationship with SP expression as a defence response to stimuli such as the masticatory function and occlusal trauma. Articles included in this review were searched in PubMed, Scopus and ISI Web of Science databases, combining the following keywords: human dentine pulp, angiogenesis, angiogenic growth factors, neuropeptides, substance P, neurogenic inflammation, dentine matrix, dentinogenesis, occlusal trauma and dental occlusion. It is concluded that human dental pulp tissue responds to occlusal trauma and masticatory function with a neurogenic inflammatory phenomenon in which SP plays an important role in the direct and indirect mechanisms of angiogenesis by the action evoked via NK1 receptors at different cells, such as fibroblasts, endothelial and inflammatory cells, leading to new blood vessel formation which are needed to stimulate mineralized tissue formation as a defence mechanism.

The influence of two reciprocating single-file and two rotary-file systems on the apical extrusion of debris and its biological relationship with symptomatic apical periodontitis. A systematic review and meta-analysis.

This systematic review and meta-analysis investigated the influence of the number of files (full-sequence rotary-file versus reciprocating single-file systems) used during root canal preparation on the apical extrusion of debris and its biological relationship with the occurrence of symptomatic apical periodontitis. An extensive literature research was carried out in the Medline, ISI Web of Science and Cochrane databases, for relevant articles with the keyword search strategy. Based on inclusion and exclusion criteria, two reviewers independently rated the quality of each study determining the level of evidence of the articles selected. The primary outcome for the meta-analysis was determined by the amount of debris extruded into the periapical tissue during root canal preparation with multiple- or single-file systems in four laboratory studies. Analysis of in vivo release of neuropeptides (SP and CGRP) after root canal preparation with single- or multiple-file systems was also carried out. Amongst the 128 articles initially found, 113 were excluded for being nonrelevant or not fulfilling the selection criteria. Another four articles were excluded after methodology evaluation. Finally, nine laboratory studies and two in vivo studies were included in the systematic review. Four of the laboratory studies were further included for meta-analysis that revealed greater debris extrusion after the use of single-file techniques when compared to multiple-file systems. Analysis of in vivo neuropeptide expression in the periodontal ligament suggests that the design of the instrument is more important than the number of files used. Both rotary and reciprocating single-file systems generate apical extrusion of debris in laboratory studies, or expression of neuropeptides in vivo. Available evidence is limited, but supports the fact that this inflammatory reaction is not influenced by the number of files but the type of movement and the instrument design.

The effect of single-file reciprocating systems on Substance P and Calcitonin gene-related peptide expression in human periodontal ligament.

AIM: To quantify the effect of two single-file reciprocating root canal preparation systems on Substance P (SP) and Calcitonin gene-related peptide (CGRP) expression in healthy human periodontal ligament (PDL). METHODOLOGY: Forty PDL samples were obtained from healthy premolars where extraction was indicated for orthodontic reasons. Prior to extraction, 20 of these premolars were divided equally in two groups, and then, root canals were prepared using one of two different single-file systems: WaveOne and Reciproc. Ten premolars were prepared with hand files and served as a positive control group. The remaining 10 premolars where extracted without treatment and served as a negative control group. All PDL samples were processed, and SP and CGRP were measured by radioimmunoassay. RESULTS: Greater SP and CGRP expression were found in the hand instrumentation group (1.220 pmol SP and 0.084 pmol CGRP per mg of PDL), followed by the WaveOne group (0.908 pmol SP and 0.046 pmol CGRP per mg of PDL) and the Reciproc group (0.511 pmol SP and 0.022 pmol CGRP per mg of PDL). The lower SP and CGRP values were associated with the intact control group (0.453 pmol SP and 0.018 pmol CGRP per mg of PDL). The Kruskal-Wallis test revealed significant differences between groups (P < 0.001). Post hoc Tukey HSD tests revealed significant differences in SP and CGRP expression between intact teeth in the control group and all the other groups (P < 0.001) except with the Reciproc group (P = 0.165 and P = 0.42 for SP and CGRP, respectively). Hand instrumentation was associated with significant differences with all the other groups (P < 0.001). Differences between the WaveOne and Reciproc groups were also significant (P < 0.001). CONCLUSION: Substance P and CGRP expression in PDL cells increased when teeth were prepared with WaveOne as well as with hand instrumentation. Reciproc maintained SP and CGRP levels in line with the negative control group.

The effect of dentine-bonding agents on substance P release in human dental pulp.

AIM: To quantify the effect of dentine-bonding agents on Substance P (SP) release in healthy human dental pulp tissue. METHODOLOGY: Forty pulp samples were obtained from healthy pre-molars where extraction was indicated for orthodontic reasons. In thirty of these pre-molars, a standardized Class V cavity preparation was performed, and teeth were divided equally into three groups: (i) Unetched-cavity control group: Class V cavities only; (ii) Experimental Group I: 'One-step' self-etch bonding agent was placed in the cavity; and (iii) Experimental Group II: 'Two-step' total-etch bonding agent was placed in the cavity. The remaining ten healthy pre-molars where extracted without treatment and served as an intact-teeth control group. SP was measured by radioimmunoassay. RESULTS: Greater SP release was found in the 'one-step' bonding agent group, followed by the 'two-step' bonding agent group and the unetched-cavity control group. The lower SP values were for the intact-teeth control group. anova showed statistically significant differences between groups (P = 0.0001). Tukey HSD post hoc tests showed statistically significant differences in SP release between the intact-teeth control group and the three other groups (P < 0.01) and between the unetched-cavity control group and the 'one-step' bonding agent group (P < 0.05). No significant difference was found between the 'two-step' bonding agent and the unetched-cavity control group. CONCLUSION: Dentine-bonding agents placed over Class V cavity preparations increased SP release. One-step dentine-bonding agents increased SP release most.

Expression of insulin-like growth factor-1 and proliferating cell nuclear antigen in human pulp cells of teeth with complete and incomplete root development.

AIM: To quantify the expression of insulin-like growth factor-1 (IGF-1) and proliferating cell nuclear antigen (PCNA) in human pulp cells of teeth with complete or incomplete root development, to support the specific role of IGF-1 in cell proliferation during tooth development and pulp reparative processes. METHODOLOGY: Twenty six pulp samples were obtained from freshly extracted human third molars, equally divided in two groups according to root development stage (complete or incomplete root development). All samples were processed and immunostained to determine the expression of IGF-1 and PCNA in pulp cells. Sections were observed with a light microscope at 80x and morphometric analyses were performed to calculate the area of PCNA and IGF-1 immunostaining using digital image software. Mann-Whitney's test was used to determine statistically significant differences between groups (P < 0.05) for each peptide and the co-expression of both. RESULTS: Expression of IGF-1 and PCNA was observed in all human pulp samples with a statistically significant higher expression in cells of pulps having complete root development (P = 0.0009). CONCLUSION: Insulin-like growth factor-1 and PCNA are expressed in human pulp cells, with a significant greater expression in pulp cells of teeth having complete root development.

Vasoactive intestinal peptide receptor expression in chronic periapical lesions.

AIM: To use radioreceptor analysis for evaluating whether vasoactive intestinal peptide (VIP) receptors are present in chronic periapical lesions and to determine whether differences in its expression are found according to the size of the lesions. METHODOLOGY: Twelve periapical lesions were obtained from teeth diagnosed with chronic apical periodontitis and indicated for endodontic surgery; they were classified according to the size of the lesion in two groups of six samples (lesion size greater or smaller than 5 mm), and then processed and labelled with (125)I-VIP. Binding sites were identified by (125)I-VIP and standard VIP competition assays. Mann-Whitney's test was used to establish statistically significant differences in the VIP receptor expression between groups. RESULTS: Vasoactive intestinal peptide receptor expression was found in all periapical lesion samples. There was a statistically significantly higher expression in periapical lesions <5 mm (P < 0.001). CONCLUSION: Vasoactive intestinal peptide receptors were expressed in chronic periapical lesions with levels inversely proportional to lesion size.

Substance P receptor expression in healthy and inflamed human pulp tissue.

AIM: To use radioreceptor analysis for comparing substance P (SP) receptor expression in human pulp tissue samples collected from teeth having a clinical diagnosis of acute irreversible pulpitis, healthy pulps and teeth with induced inflammation. METHODOLOGY: Five pulp samples were obtained from teeth having a clinical diagnosis of acute irreversible pulpitis. Another 10 pulp samples were obtained from healthy premolars where extraction was indicated for orthodontic purposes. In five of these premolars inflammation was induced prior to pulp collection. All of the samples were processed and labelled with 125I-SP. Binding sites were identified by 125I-SP and standard SP competition assays. Kruskal-Wallis and Mann-Whitney (post-hoc) tests were used to establish statistically significant differences between the groups. RESULTS: Substance P receptor expression was found in all human pulp tissue samples. Most receptors were found in the group of pulps from teeth having a clinical diagnosis of acute irreversible pulpitis, followed by the group of pulps having induced inflammation. The least number of receptors was expressed in the group of healthy pulps. Statistical analysis revealed significant differences between the group of healthy pulp and both inflamed pulp groups (P < 0.01). CONCLUSION: Substance P receptor expression in human pulp tissue is significantly increased during inflammatory phenomena such as acute irreversible pulpitis.

Quantification of neuropeptides (calcitonin gene-related peptide, substance P, neurokinin A, neuropeptide Y and vasoactive intestinal polypeptide) expressed in healthy and inflamed human dental pulp.

AIM: To quantify the expression of calcitonin gene-related peptide (CGRP), substance P (SP), neurokinin A (NKA), neuropeptide Y (NPY) and vasoactive intestinal polypeptide (VIP) in healthy and inflamed human dental pulp tissue. METHODOLOGY: Six pulp samples were obtained from teeth having a clinical diagnosis of acute irreversible pulpitis. Another 12 pulp samples were obtained from premolars where extraction was indicated for orthodontic purposes. In six of these premolar teeth inflammation was induced by mechanical pulp exposure prior to sample collection. All samples were processed and 125I-labelled; neuropeptides were quantified by competition assays. ANOVA and Mann-Whitney's (post hoc) tests were used to establish statistically significant differences between the groups. RESULTS: Expression of five neuropeptides was found in all human pulp samples. Statistical analysis revealed a significantly higher (P < 0.05) expression of CGRP, SP, NKA and NPY in both inflammatory conditions compared with healthy pulp control values. VIP expression remained stable during the inflammatory conditions. CONCLUSION: Expression of CGRP, SP and NKA released from C-fibres and NPY released from sympathetic fibres is significantly higher in the inflamed human pulp compared with healthy pulp. Expression of VIP released from parasympathetic fibres is not increased during the inflammatory conditions of human dental pulp.

Calcitonin gene-related peptide receptor expression in healthy and inflamed human pulp tissue.

AIM: To use radioreceptor analysis for comparing calcitonin gene-related peptide (CGRP) receptor expression in human pulp tissue samples collected from teeth having a clinical diagnosis of acute irreversible pulpitis, healthy pulps and teeth with induced inflammation. METHODOLOGY: Six pulp samples were obtained from teeth having a clinical diagnosis of acute irreversible pulpitis. Another eight pulp samples were obtained from healthy premolars where extraction was indicated for orthodontic purposes. In four of these premolars, inflammation was induced prior to pulp collection. All the samples were processed and labelled with 125I-CGRP. Binding sites were identified by 125I-CGRP and standard CGRP competition assays. RESULTS: CGRP receptor expression was found in all human pulp tissue samples. Most receptors were found in the group of pulps from teeth having a clinical diagnosis of acute irreversible pulpitis, followed by the group of pulps having induced inflammation. The least number of receptors was expressed in the group of healthy pulps. The Kruskal-Wallis and Mann-Whitney (post-hoc) tests showed statistically significant differences between the groups (P < 0.05). CONCLUSION: CGRP receptor expression in human pulp tissue is significantly increased during inflammatory phenomena such as acute irreversible pulpitis.

The effect of capsaicin on substance P expression in pulp tissue inflammation.

AIM: To evaluate the effect of capsaicin on substance P (SP) expression during induced inflammation in rat pulp tissue. METHODOLOGY: Radioimmunoanalysis was used to measure SP levels in 36 mandibular molar pulps taken from six Wistar rats. Twelve samples were obtained from healthy pulps and used as negative control group. Another 12 samples were obtained after inducing inflammation with mechanical pulp exposure; these were used as the positive control group. Capsaicin was infiltrated into the inferior dental nerve in the experimental group and 12 samples were obtained after mechanical pulp exposure. RESULTS: The lowest SP expression was found in mechanically exposed pulps where capsaicin pretreatment had been carried out (0.028 ng mL(-1)), followed by healthy pulps (0.302 ng mL(-1)). The highest SP expression was found in mechanically exposed pulps with no capsaicin pretreatment (124 ng mL(-1)). The Kruskal-Wallis test showed statistically significant differences between the groups (P < 0.001). CONCLUSION: Inferior dental nerve infiltration with capsaicin reduces SP expression in dental pulp tissue in rats.