CSF Single-cell Sequencing and Proteomics of Chronic Postsurgical Pain in Patients With Lower Limb Fractures

Study Overview
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Brief Summary
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The purpose of this study is to assess the proteomics and transcriptomic differences between pain-free control subjects and patients with chronic postoperative pain through single-cell sequencing technology.

Detailed Description
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Chronic pain,one of the most frequent causes for patients to seek medical care,is a recognized health problem.Chronic postsurgical pain (CPSP), commonly defined as pain that develops after a surgical procedure and persists at least 3 months, constitutes a widely underdiagnosed and often poorly treated medical problem affecting 10-50% of all postsurgical patients. According to the reports,in the United States alone,1.9 million persons abused or were dependent on prescription opioid analgesics for chronic pain in 2013,contributing to one of the worst public health crises the developed world has recently faced.Here,open reduction and internal fixation of lower limb fractures ,what we focus, is a common surgical procedure in orthopedics and microscopic hand and foot surgery.The removal of the intramedullary nail can relieve anterior knee pain, but in a substantial number of patients, pain persists after nail removal.The most painful daily activities are kneeling and squatting.Therefore, it is increasingly important and urgent to solve the postoperative chronic pain of patients with lower extremity fractures surgery. The underlying biology of chronic postoperative pain and genetic heritability is complex and not yet fully understood . A common feature of CPSP is that the painful sensations change from the familiar acute postoperative pain to a complex pain syndrome with nociplastic characteristics,neuropathic characteristics, or both. Preclinical studies have revealed that neuroinflammation is one of pathological hallmarks of CPSP. The transition from acute to chronic pain starts early within the first 2 weeks after nociception by peripheral and central inflammatory processes and activation of spinal glial cells.Repetitive nociception resulting from prolonged inflammatory and neuropathic responses to noxious stimuli causes a cascade of biochemical and structural changes to various pain pathways resulting in sensitization of the peripheral and CNS. Cytokines and neurotrophic factors have been identified as pivotal mediators involved in neuroimmune activation pathways and cascades in various preclinical chronic pain models. The brain is surrounded by the meninges, a membranous covering that contains the cerebrospinal fluid (CSF).Its circulatory system transmits peripheral stimulation to the cerebrum, and central signal is transmitted to the dorsal root ganglion. CSF ,as a bridge between peripheral and central, contains a tightly regulated immune system and metabolitites and reflects the inflammatory processes and activation of spinal glial cells. However, knowledge is lacking about how CSF contains are altered with pain.We thus hypothesized that comparing the CSF protemics and immune transcriptomes associated with pain-free control subjects and patients with chronic postoperative pain would provide insights into the pathophysiology of CPSP. Here,a single-cell transcriptomic resource exploring the cerebrospinal fluid immune system and proteomics of patients with postoperative chronic pain will uncover the key molecules and pathways of CPSP.

Official Title
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Cerebrospinal Fluid Single-cell Sequencing and Proteomics Analysis of the Chronic Postsurgical Pain in Patients Following Open Reduction Internal Fixation of Lower Limb Fractures

Conditions
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Chronic Postsurgical Pain

Participation Criteria
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Eligibility Criteria
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Inclusion Criteria: ASA physical status 1 to 3 (males and females) Undergone open reduction and internal fixation of the lower limbs and plan to take out the internal fixation under spinal anesthesia Exclusion Criteria: Absence of written consent Contraindication to regional anaesthesia Known dementia at time of operation History or opioid abuse Unexpected conversion of general anesthesia or surgery is not carried out
Sexes Eligible for Study
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All

Accepts Healthy Volunteers
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No

Study Plan
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How is the study designed?
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Arms and Interventions

| Participant Group/Arm | Intervention/Treatment |
| --- | --- |
| Pain-free control subjects<br>According to the 11th Revision of International Classification of Diseases (ICD-11) of International Association for the Study of Pain(IASP) in 2019, diagnose whether patients have CPSP . Patients with no postoperative chronic pain after open reduction and internal fixation of lower limb fractures. | |
| Patients with chronic postsurgical pain<br>According to the 11th Revision of International Classification of Diseases (ICD-11) of International Association for the Study of Pain(IASP) in 2019, diagnose whether patients have CPSP . Patients with postsurgical chronic pain after open reduction and internal fixation of lower limb fractures. | |

What is the study measuring?
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Primary Outcome Measures

| Outcome Measure | Measure Description | Time Frame |
| --- | --- | --- |
| Significantly different proteomics and transcriptomes between Chronic Postsurgical Pain and pain-free subjects | Differences in the proteins in the fluid around the lumbar, between Chronic Postsurgical Pain and Pain-free subjects.，will be quantified using Mass spectrometry，for: neural cell adhesion molecule L1, complement C4-A, lysozyme C, receptor-type tyrosine-protein phosphatase zeta, apolipoprotein D, alpha-1-antichymotrypsin, granulins, calcium/calmodulin-dependent protein kinase type II subunit alpha, mast/stem cell growth factor receptor Kit, prolow-density lipoprotein receptor-related protein 1 and so on. These proteins may identify the disease and define its mechanism. | Up to 3 months after fixation | 
| Inflammatory cytokines in Chronic Postsurgical Pain patients | Biomarkers from CSF will be quantified using Meso Scale Discovery multiplex kit (K15210D), for: CRP, Eotaxin, Eotaxin-3, FGF (basic), ICAM-1, IFN-γ, IL-1α, IL-1β, IL-2, IL-4, IL-5, IL-6, IL-7, IL-8, IL-10, IL-12/IL-23p40, IL-13, IL-15, IL-16, IL-17A, IP-10, MCP-1, MCP-4, MDC, MIP-1α, MIP-1β, PlGF, SAA, TARC, Tie-2, TNF-α, TNF-β, VCAM-1, VEGF-A, VEGF-C, VEGF-D, VEGFR-1/Flt-1. The levels of these biomarkers will be compared between the group between CPSP and pain-free subjects. | Up to 3 months after fixation surgery | 

 Terms related to the study
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Keywords Provided by Centre Hospitalier Valida
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Chronic Postsurgical pain, Single-cell sequencing, Cerebrospinal Fluid