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Voltage-gated sodium channels, especially Nav1.7 encoded by the SCN9A gene, are key regulators of nociceptive transmission. Upregulation of SCN9A has been associated with increased neuronal excitability and heightened pain perception. In parallel, inflammatory cytokines such as interleukin-6 (IL-6), tumor necrosis factor-alpha (TNF-α), and interleukin-1 beta (IL-1β) are known to sensitize peripheral nociceptors and reduce the efficacy of local anesthetics by modifying tissue environment and ion channel activity. However, the combined influence of SCN9A expression and inflammatory cytokines on anesthetic success in SIP has not been fully elucidated.
This prospective case-control study aims to evaluate the association between SCN9A gene expression and inflammatory cytokine levels with the clinical success of IANB in patients with SIP affecting mandibular molars. Approximately 90-100 patients will be recruited and categorized into two groups based on anesthetic outcome: successful anesthesia and failed anesthesia. All patients will receive a standardized IANB using 2% lidocaine with 1:100,000 epinephrine. Anesthetic success will be determined based on the absence of pain during access cavity preparation and instrumentation.
Following access and pulp extirpation, pulpal tissue samples will be collected. SCN9A gene expression will be assessed using quantitative real-time polymerase chain reaction (RT-qPCR), with relative expression calculated using the 2^-ΔΔCt method. Inflammatory cytokine levels (IL-6, TNF-α, IL-1β) will be quantified using enzyme-linked immunosorbent assay (ELISA).
The primary outcome will be the difference in SCN9A expression between failed and successful anesthesia groups. Secondary outcomes will include comparison of cytokine levels and evaluation of correlations between SCN9A expression and inflammatory markers. Statistical analysis will include group comparisons, correlation analysis, logistic regression, and receiver operating characteristic (ROC) curve analysis to assess the predictive value of these biomarkers.
The present study is designed as a prospective case-control investigation to assess the association between SCN9A gene expression and levels of key inflammatory cytokines with the clinical success of IANB. A total of approximately 100 patients diagnosed with SIP in mandibular molars will be recruited and categorized into two groups based on anesthetic outcome: successful anesthesia and failed anesthesia. Standardized IANB will be administered using 2% lidocaine with 1:100,000 epinephrine, and anesthetic success will be determined based on absence of pain during access cavity preparation and instrumentation.
Following access cavity preparation and pulp extirpation, biological samples will be collected. Pulpal tissue samples will be used for RNA extraction and subsequent quantitative real-time polymerase chain reaction (RT-qPCR) analysis to assess SCN9A gene expression. Relative expression levels will be calculated using the 2^-ΔΔCt method with appropriate housekeeping genes. In parallel, inflammatory cytokine levels (IL-6, TNF-α, IL-1β) will be quantified using enzyme-linked immunosorbent assay (ELISA) from pulpal tissue homogenates or gingival crevicular fluid, depending on feasibility.
The primary outcome of the study will be the difference in SCN9A expression between failed and successful anesthesia groups. Secondary outcomes will include comparison of cytokine levels between groups and evaluation of correlations between SCN9A expression and inflammatory markers. Data will be analyzed using appropriate statistical tests, including independent t-tests or non-parametric equivalents, correlation analysis, and logistic regression modeling. Additionally, receiver operating characteristic (ROC) curve analysis may be performed to assess the predictive value of these biomarkers for anesthetic failure.
This study aims to provide mechanistic insights into anesthetic failure in SIP by integrating molecular and inflammatory pathways. The findings may contribute to the development of predictive biomarkers and targeted therapeutic strategies to improve anesthetic success in endodontic practice.
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| Label | Type | Description | Intervention Names |
|---|---|---|---|
| IANB Success (Adequate Anesthesia) Group | This cohort will include patients with symptomatic irreversible pulpitis in mandibular molars who achieve successful pulpal anesthesia following administration of a standardized inferior alveolar nerve block (IANB) using 2% lidocaine with 1:100,000 epinephrine. Anesthetic success will be defined as the absence of pain (no or mild pain) during access cavity preparation and initial instrumentation without the need for any supplementary anesthetic techniques. Following confirmation of anesthesia, access cavity preparation and pulp extirpation will be performed, and pulpal tissue samples will be collected for analysis of SCN9A gene expression (RT-qPCR) and inflammatory cytokine levels (IL-6, TNF-α, IL-1β) using ELISA. |
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| IANB Failure (Inadequate Anesthesia) Group | This cohort will include patients with symptomatic irreversible pulpitis in mandibular molars who experience inadequate pulpal anesthesia following administration of a standardized inferior alveolar nerve block (IANB) using 2% lidocaine with 1:100,000 epinephrine. Anesthetic failure will be defined as the presence of moderate to severe pain during access cavity preparation or instrumentation, necessitating the use of supplementary anesthetic techniques (e.g., intraligamentary or intrapulpal injections). Pulpal tissue samples will be collected after access cavity preparation and pulp extirpation for analysis of SCN9A gene expression (RT-qPCR) and inflammatory cytokines (IL-6, TNF-α, IL-1β) using ELISA. |
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| Name | Type | Description | Arm Group Labels | Other Names |
|---|---|---|---|---|
| Inferior Alveolar Nerve Block | Procedure | All participants will receive a standardized inferior alveolar nerve block (IANB) administered using 2% lidocaine with 1:80,000 epinephrine. The injection will be performed using a conventional Halsted technique with a 27-gauge long needle under strict aseptic conditions. The needle will be inserted at the pterygomandibular raphe region, advancing until bony contact is achieved near the mandibular foramen. Following negative aspiration, approximately 1.8 mL of anesthetic solution will be deposited slowly over 60-90 seconds. Lip numbness will be assessed after 10-15 minutes to confirm nerve block onset. No additional anesthetic techniques will be used prior to the assessment of primary anesthetic success. Endodontic access cavity preparation will then be initiated, and pain response during access and initial instrumentation will be recorded using a standardized pain scale. Anesthetic success or failure will be determined based on the patient's pain response, as per predefined criteria. |
| Measure | Description | Time Frame |
|---|---|---|
| Success of Inferior Alveolar Nerve Block (IANB) | The primary outcome will be the clinical success of the inferior alveolar nerve block (IANB), assessed during endodontic access cavity preparation and initial instrumentation. Anesthetic success will be defined as the absence of pain or the presence of only mild pain that does not require any supplementary anesthetic intervention. Anesthetic failure will be defined as the presence of moderate to severe pain necessitating additional anesthesia (e.g., intraligamentary or intrapulpal injection). | Assessed 15 minutes after administration of IANB, during access cavity preparation and initial instrumentation at the same visit |
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Inclusion Criteria:
Exclusion Criteria:
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The study population will consist of adult patients presenting to the postgraduate endodontic clinic with symptomatic irreversible pulpitis in mandibular first or second molars requiring root canal treatment. Eligible participants will be systemically healthy (ASA I or II), aged between 18 and 60 years, and will report moderate to severe preoperative pain. Diagnosis will be established based on clinical findings, including spontaneous pain, prolonged response to cold testing, and absence of periapical pathology requiring surgical intervention.
| Name | Role | Phone | Extension | |
|---|---|---|---|---|
| Vivek Aggarwal, MDS | Contact | 01126983646 | vaggarwal@jmi.ac.in |
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| Facility | Status | City | State | ZIP | Country | Contacts |
|---|---|---|---|---|---|---|
| Faculty of Dentistry | Recruiting | New Delhi | National Capital Territory of Delhi | 110025 | India |
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The primary biospecimen will be vital pulpal tissue, obtained aseptically immediately after access cavity preparation and pulp extirpation. The collected pulp tissue will be divided into two portions: one portion will be preserved in RNA stabilization solution for subsequent RNA extraction and gene expression analysis of SCN9A; the second portion will be processed for protein extraction and stored appropriately for the estimation of inflammatory cytokines (IL-6, TNF-α, IL-1β) using enzyme-linked immunosorbent assay (ELISA).
All samples will be de-identified and labeled with unique study codes to ensure confidentiality. Biospecimens will be stored under controlled laboratory conditions and used exclusively for the purposes of this study. No genetic will be used for purposes beyond those outlined in the study protocol without additional ethical approval.
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