JP2017187492A - Pain evaluation method - Google Patents

Abstract

PROBLEM TO BE SOLVED: To provide objective indicators which allow for eliminating psychosocial effect that modifies pain complaints and evaluating presence/absence and severity of neuropathic pain according to the nervous condition.SOLUTION: A method of acquiring data for objective pain evaluation involves detecting lysophosphatidic acid (LPA) and/or lysophosphatidylcholine (LPC) in a sample derived from human cerebrospinal fluid.SELECTED DRAWING: Figure 1-1

Description

  The present invention relates to an objective evaluation method for pain such as neuropathic pain.

  Pain is broadly divided into nociceptive pain caused by inflammation and stimulation, neuropathic pain caused by nerve injury and compression, and psychogenic pain caused by psychological and social factors. . Among them, neuropathic pain is pain caused by lesions or dysfunction in the central nervous system or peripheral nervous system, and is caused by nerve damage such as trauma, tumor, viral infection, metabolic disorder, spinal canal stenosis, intervertebral disc herniation, etc. It is caused by nerve damage.

  Neuropathic pain is estimated to have a prevalence of 6-7% in developed countries, with approximately 50% of patients likely to become chronic with a disease duration of 3 years or more, and is the most severe chronic pain disorder It is expensive. Most of the neuropathic pain is often caused by age-related changes in the musculoskeletal system, particularly the spine, and it is estimated that the number of patients will increase further in the future as the super-aged society progresses.

  A problem in the treatment of neuropathic pain is the lack of methods for objectively assessing pain. That is, pain is a subjective measure, and there are patients who complain of pain and those who are not, objectively the same sign. Pain may become chronic even after tissue damage, such as trauma, has been improved, making it difficult for patients to perform daily living activities (ADL) and reduce quality of life (QOL). . In order to appropriately treat such pain, a test method for objectively determining pain symptoms is required, and the development of biomarkers related to neuropathic pain is required in the clinic. Yes.

  Now, in a mouse experimental model, lysophosphatidic acid (LPA) has been reported as a causative substance that causes neuropathic pain (Non-Patent Documents 1 to 3). These reports have experimentally confirmed the induction of pain by LPA subarachnoid administration and the disappearance of pain in LPA receptor knockout mice.

  On the other hand, interstitial cystitis is a relatively rare intractable disease mainly having frequent urination, urgency, and bladder pain. Unlike cystitis due to bacterial infection, there is no abnormality in urinary findings or urine culture. For the diagnosis of interstitial cystitis, cystoscopy and hydraulic dilatation under anesthesia are performed. Treatment includes drug therapy, diet therapy, etc., but the effects are limited. In severe cases, surgical treatment such as bladder enlargement and cystectomy is performed. Surgical treatment is predicated on reliable diagnosis. However, endoscopic findings lack objectivity, and it is difficult to make a definitive diagnosis unless the doctor has experience in many cases.

Nature Medicine, Vol.10, No.7, 712-718, 2004 Pharmacological Journal of Japan, 127, 161-165, 2006 Biol. Pharma. Bull., 34 (8) 1154-1158, 2011

In addition to neuropathic pain, pain is a subjective symptom, and it is very difficult to objectively evaluate the presence and severity of pain, and it is difficult to elucidate the cause of pain itself There are also many. However, it is an unbearable pain for patients, and it is required to eliminate or reduce the pain as early as possible.
At present, neuropathic pain is generally evaluated based on a patient's subjective complaint using a neuropathic pain evaluation questionnaire (Neuropathic Pain Symptom Inventory, NPSI) or the like. Therefore, it is difficult to obtain an evaluation result that excludes psychosocial effects that modify pain complaints. In many cases, more invasive treatments such as drug treatment and surgery based on subjective complaints do not produce the expected effects, so the ambiguity in pain assessment is excluded and the presence or absence of neuropathic pain. And an index to assess severity was needed.
Interstitial cystitis has similar symptoms to infectious cystitis, overactive bladder, bladder cancer and the like, and is often misdiagnosed. In addition, cystoscopy under water pressure expansion is necessary for diagnosis, and considerable experience is required to determine specific endoscopic findings. Therefore, in order to start treatment of interstitial cystitis at an early stage, a method capable of objectively and accurately making a diagnosis is desired.

  Various lysophospholipids exist in the living body, and some of them are known to have a strong bioactive action. As described above, it is reported that LPA may be the causative agent of pain in experiments in mice. However, in human patients, there is still a clear report on the relationship between pain actually recognized and LPA in the body. Has not been. In addition, LPA and other glycerolysophospholipids are known to have multiple molecular species due to their constituent fatty acid chains, and LPA has increased biological activity depending on the molecular species and disease. It has been reported that molecular species to be biased are biased. However, none of the molecular species of lysophospholipids are important in relation to neuropathic pain, and no lysophospholipids other than LPA have been reported.

The present inventors can specifically examine whether or not spinal surgery can be applied, and LPA and lysophosphatidylcholine (Lysophosphatidylcholine) in cerebrospinal fluid for patients with lumbar spinal canal stenosis considered to have very little psychosocial modification. , LPC) and lysophospholipids were simultaneously determined for each molecular species having different fatty acids. As a result, in addition to LPA, a high correlation between cerebrospinal fluid concentration and pain was also found for LPC, which has not been reported in basic research in the past. Furthermore, in patients with subjective symptoms of pain, not only significantly high concentrations of LPA and LPC were detected in the cerebrospinal fluid, but also to a degree that varies depending on the type of molecular species of LPA and LPC. It became clear.
Furthermore, by quantifying the concentration of LPA and LPC molecular species in the cerebrospinal fluid, we found that it is possible to objectively evaluate pain caused by other diseases such as interstitial cystitis. It was.
Based on these findings, the present inventors have completed the present invention.

That is, the present invention provides the following.
1. A method for acquiring data for objective evaluation of pain, comprising detecting lysophosphatidic acid (LPA) and / or lysophosphatidylcholine (LPC) in a sample derived from human cerebrospinal fluid.
2. 2. The method according to 1 above, wherein the detection of LPA and / or LPC is detection for each molecular species.
3. 3. The method according to 1 or 2 above, wherein the pain is neuropathic pain, and the objective evaluation is determination of the presence and / or severity of neuropathic pain.
4). 4. The method according to any one of 1 to 3 above, wherein the LPA is at least one selected from the total LPA, LPA_16_0, LPA_18_0, LPA_18_1, LPA_18_2, LPA_20_4, and LPA_22_6.
5. 5. The method according to 4 above, wherein at least LPA_16_0 and / or LPA_18_1 is detected.
6). 6. The method according to any one of 1 to 5 above, wherein the LPC is at least one selected from LPC_14_0, LPC_16_0, LPC_16_1, LPC_18_0, LPC_18_1, LPC_18_2, LPC_20_4, LPC_20_5, and LPC_22_6.
7). 7. The method according to 6 above, wherein at least LPC_20_4 and / or LPC_22_6 is detected.
8). 8. The method according to any one of 1 to 7 above, wherein at least LPA_16_0, LPA_18_1, LPC_20_4, and LPC_22_6 are detected.
9. 9. The method according to any one of 1 to 8 above, which is detected by an enzymatic method or mass spectrometry.
10. 10. The method according to any one of 1 to 9, further comprising comparing the detection result with a reference value for determining the presence and / or severity of neuropathic pain.
11. Based on the results of the evaluation, the neuropathic pain assessment questionnaire (Neuropathic Pain Symptom Inventory, NPSI) and / or the Zurich Claudication Questionnaire (ZCQ), pain assessment, morphological severity assessment by nuclear magnetic resonance imaging, The method according to any one of 1 to 10 above, wherein the data is used for objective evaluation in combination with nerve damage evaluation in which phosphorylated neurophilament heavy chain (pNFH) is determined as a marker.
12 A diagnostic marker for determining the presence and / or severity of neuropathic pain, which includes an increase in the amount of LPA and / or LPC in cerebrospinal fluid as an indicator.
13. A kit for determining the presence and / or severity of neuropathic pain, comprising a reagent for quantitative detection of LPA and / or LPC in cerebrospinal fluid, reference value information in a control sample or a control sample.
14 3. The method according to 1 or 2 above, wherein the pain is pain caused by interstitial cystitis.
15. 15. The method according to 14 above, wherein the LPA is one or more selected from LPA_16_0, LPA_18_1, and LPA_18_2.
16. 16. The method according to 15 above, wherein at least LPA_16_0 and / or LPA_18_2 is detected.
17. 17. The method according to any one of 14 to 16, wherein the LPC is one or more selected from LPC_16_0, LPC_16_1, LPC_18_0, LPC_18_1, LPC_18_2, and LPC_22_6.
18. 18. The method according to 17 above, wherein at least LPC_16_0, LPC_18_1 and / or LPC_22_6 is detected.
19. 19. The method according to any one of 14 to 18, further comprising comparing the detection result with a reference value for determining pain due to interstitial cystitis.
20. A diagnostic marker for the determination of pain caused by interstitial cystitis, which includes, as an index, a reduction in the amount of LPA and / or an increase in the amount of LPC in cerebrospinal fluid.
21. A kit for the determination of pain due to interstitial cystitis, comprising a reagent for quantitative detection of LPA and / or LPC in cerebrospinal fluid, reference value information in a control sample or a control sample.

  The present invention can enable objective evaluation of neuropathic pain, which has been difficult in the past. By combining the method of the present invention with conventional pain evaluation methods such as NPSI and ZCQ, MRI image diagnosis, etc., the patient's condition can be determined more accurately, leading to appropriate treatment. In addition, the method of the present invention can be applied to all pains other than neuropathic pain.

The amount of lysophosphatidic acid (LPA) in the samples from the patient group and the control group is shown respectively. a: total LPA; b: LPA_16_0; c: LPA_18_0; d: LPA_18_1. The amount of lysophosphatidic acid (LPA) in the samples from the patient group and the control group is shown respectively. e: LPA_18_2; f: LPA_20_4. The amounts of lysophosphatidylcholine (LPC) in the samples from the patient group and the control group are shown respectively. a: total LPC; b: LPC_14_0; c: LPC_16_0; d: LPC_16_1. The amounts of lysophosphatidylcholine (LPC) in the samples from the patient group and the control group are shown respectively. e: LPC_18_0; f: LPC_18_1; g: LPC_18_2; h: LPC_20_4. The amount of lysophosphatidylcholine (LPC) in samples from the patient group and the control group is shown. i: LPC_22_6. The amount of lysophosphatidic acid (LPA) in the sample derived from the patient group judged to be mild and severe by the score in NPSI is shown, respectively. a: LPA_18_2; b: LPA_20_4; c: LPA_22_6. The amounts of lysophosphatidylcholine (LPC) in samples from patient groups determined to be mild and severe by NPSI score are shown. a: LPC_14_0; b: LPC_16_0; c: LPC_16_1; d: LPC_18_0. The amounts of lysophosphatidylcholine (LPC) in samples from patient groups determined to be mild and severe by NPSI score are shown. e: LPC_18_1; f: LPC_18_2; g: LPC_20_4; h: LPC_20_5. The amount of lysophosphatidylcholine (LPC) in a sample from a group of patients determined to be mild and severe by the score in NPSI is shown. i: LPC_22_6. The amount of lysophosphatidic acid (LPA) in the sample derived from a patient group determined to be mild and severe by the score in ZCQ is shown. a: LPA_18_1; b: LPA_18_2; c: LPA_20_4; d: LPA_22_6. The amounts of lysophosphatidylcholine (LPC) in samples from patient groups determined to be mild and severe by ZCQ score are shown. a: LPC_14_0; b: LPC_16_0; c: LPC_16_1; d: LPC_18_0. The amounts of lysophosphatidylcholine (LPC) in samples from patient groups determined to be mild and severe by ZCQ score are shown. e: LPC_18_1; f: LPC_18_2; g: LPC_20_4; h: LPC_20_5. The amount of lysophosphatidylcholine (LPC) in a sample from a patient group determined to be mild and severe in the score in ZCQ is shown. i: LPC_22_6. The amounts of lysophosphatidic acid (LPA) in samples from patient groups determined to be mild (grade A-B) and severe (grade C-D) using diagnostic imaging by the method of Schizas et al. Are shown, respectively. a: LPA_16_0; b: LPA_18_0; c: LPA_18_1; d: LPA_20_4. The amounts of lysophosphatidylcholine (LPC) in samples from patient groups determined to be mild (grade A-B) and severe (grade C-D) using image diagnosis by the method of Schizas et al. Are shown. a: LPC_14_0; b: LPC_16_0; c: LPC_16_1; d: LPC_18_0. The amounts of lysophosphatidylcholine (LPC) in samples from patient groups determined to be mild (grade A-B) and severe (grade C-D) using image diagnosis by the method of Schizas et al. Are shown. e: LPC_18_1; f: LPC_18_2; g: LPC_20_4; h: LPC_22_6.

  The present invention provides a method for acquiring data for objective evaluation of pain, characterized by detecting lysophosphatidic acid (LPA) and / or lysophosphatidylcholine (LPC) in a sample derived from human cerebrospinal fluid. To do. The method of the present invention can be, for example, a method for supporting / assisting the diagnosis of the presence and / or severity of neuropathic pain. The method of the present invention may be a method for supporting / assisting the diagnosis of interstitial cystitis.

  Cerebrospinal fluid tests are commonly performed in clinical settings for the diagnosis of various diseases, and a small amount of cerebrospinal fluid collected before performing myelography is directly used as a “sample derived from human cerebrospinal fluid”. Can be used. Since the method of the present invention is performed by obtaining such a sample, in this specification, the subject subject may be described as “patient” regardless of the presence or absence of symptoms.

  Phospholipids are amphipathic lipids constituting various biological cell membranes, and are classified into glycerophospholipids having glycerin as a skeleton and sphingophospholipids having sphingosine as a skeleton. The types of glycerophospholipids that have a structure in which two fatty acids and phosphoric acid are bonded to glycerin include phosphatidic acid (PA), phosphatidylcholine (PC), phosphatidylethanolamine (PE), phosphatidylinositol (PI), phosphatidylserine. (PS) etc. are known. A product obtained by removing one fatty acid from the glycerophospholipid by hydrolysis is called lysophospholipid. On the other hand, sphingosine 1-phosphate is an example of a lysophospholipid derived from sphingophospholipid.

  In the lysophospholipid derived from glycerophospholipid, the fatty acid remaining without being hydrolyzed is ester-bonded at the 1-position or 2-position of the glycerin skeleton, and these are respectively 1-acyl-2-lyso and 2-acyl-1- Although called a lyso form, this fatty acid is known to easily undergo intramolecular acyl transfer. A method for analyzing each of these isomers is also known (M. Okudaira et al., J. Lipid Res., 2014, 55: 2178-2192). In the present invention, these isomers are identified. Not intended.

  In this specification, the notation of lysophospholipid is described as follows according to the type of lysophospholipid (LPA, LPC, lysophosphatidylinositol, LPI) and the type of fatty acid contained in lysophospholipid.

  For example, LPA containing myristic acid as a fatty acid is referred to as “LPA — 14 — 0” in this specification. Similarly, LPC containing palmitic acid as a fatty acid is described as “LPC — 16 — 0”. In addition, LPI containing stearic acid as a fatty acid is described as “LPI — 18 — 0”. The first numbers such as “14”, “16”, “18” indicate the number of carbon atoms in the fatty acid chain, and the second numbers “0”, “1” and “2” indicate the number of double bonds in the fatty acid chain. Indicates.

  As used herein, “neuropathic pain” is pain resulting from a lesion or dysfunction in the central or peripheral nervous system, such as cervical spinal stenosis, lumbar spinal stenosis, cervical spinal cord injury, Examples include peripheral nerve damage.

  The method of the present invention is characterized by detecting LPA and / or LPC in a sample derived from human cerebrospinal fluid. In addition to LPA, the present inventors not only found an association between the content of lysophospholipids in human cerebrospinal fluid and neuropathic pain, but also a correlation between LPC and pain that has not been reported so far. I also found. In addition, the present inventors found that the concentration of LPI in the sample also showed a correlation with neuropathic pain, but sphingosine monophosphate was not correlated.

  Thus, in one embodiment, the method of the invention detects LPA in a sample. In one embodiment, the method of the invention detects LPC in a sample. In one embodiment, the method of the present invention detects LPA and LPC in a sample. LPA and LPC may be detected as total LPA and total LPC, respectively. Alternatively, LPA and LPC may be detected by detecting one or more specific molecular species for each molecular species.

  The method of the present invention enables objective assessment of neuropathic pain. As an objective evaluation, it is possible to determine the presence and / or severity of neuropathic pain.

  The presence or absence of neuropathic pain refers to the presence or absence of pain when an objective evaluation is performed excluding psychosocial effects. The severity of neuropathic pain refers to pain assessment using existing assessment methods such as the Neuropathic Pain Symptom Inventory (NPSI) and / or the Zurich Claudication Questionnaire (ZCQ), nuclear As determined by morphological severity assessment by magnetic resonance imaging, it is a scale that gives a judgment index as to whether or not treatment such as surgery is necessary, for example, two groups of "severe" and "mild" Can be divided into

  In one embodiment of the present invention, the LPA to be detected is the total LPA or one or more selected from LPA_16_0, LPA_18_0, LPA_18_1, LPA_18_2, LPA_20_4, and LPA_22_6, that is, one, two, three, four , 5 or 6 types. Although not limited, it is preferable to detect at least LPA_16_0 and / or LPA_18_1 among the above-described molecular species.

  In one embodiment of the present invention, the detection target LPC is one or more selected from LPC_14_0, LPC_16_0, LPC_16_1, LPC_18_0, LPC_18_1, LPC_18_2, LPC_20_4, LPC_20_5, and LPC_22_6, that is, one, two, or three types. There are 4, 5, 6, 7, 8, or 9 species. Although it does not limit, it is suitable to detect at least LPC_20_4 and / or LPC_22_6 among the above-mentioned molecular species.

  In a preferred embodiment of the present invention, at least LPA_16_0, LPA_18_1, LPC_20_4, and LPC_22_6 are detected.

  The methods of the present invention are not intended to detect LPA and / or LPC present at relatively high concentrations, and are objectively recognized as having neuropathic pain or severe The present invention provides a method for objectively determining neuropathic pain by finding LPA and / or LPC that can be significantly distinguished from control subjects that are not, and using this as a marker.

  Thus, the method of the present invention can selectively detect molecular species that produce more significant results in patients with and without neuropathic pain, or patients with severe and mild pain. it can.

  For example, molecular species particularly preferable as a marker for objective evaluation of the presence or absence of pain include, but are not limited to, LPA_16_0, LPA_18_1, LPC_20_4, and LPC_22_6.

  Markers for objective assessment of pain severity include LPA_16_0, LPA_18_0, LPA_18_1, LPA_18_2, LPA_20_4, LPA_22_6, LPC_14_0, LPC_16_0, LPC_16_1, LPC_18_0, LPC_18_1, LPC_18_2, LPC_18_2, PC, Can be used.

  The molecular species to be detected may be one or more of the above. However, the reliability (sensitivity and specificity) of the detection result can be further improved by combining a plurality of molecular species.

  Further, depending on the detection method, it may be advantageous to select one having a relatively high content in spinal fluid. In such a case, it is preferable to detect the molecular species including LPA_16_0 and LPA_18_1 for LPA and LPC_16_0, LPC_18_0, LPC_18_1 and the like for LPC. In addition, it may be clinically useful to detect total LPA and / or total LPC. The concentration of total LPA in the cerebrospinal fluid is in the range of 0.01 to 1.09 μM, and the concentration of total LPC is 0 to 1 μM. The LPA and / or LPC to be detected is preferably at a concentration of 0.01 μM or more in the sample.

  The detection and quantification of LPA and LPC in a sample in the method of the present invention is not limited, for example, by enzymatic method or mass spectrometry, particularly LC-MS / MS combining liquid chromatography and tandem mass spectrometry. It can be carried out.

The enzymatic method can be performed by applying a technique known in the art, and is not particularly limited. For example, for detection of LPA, fatty acid is removed from LPA by the action of lysophospholipase. The resulting glycerol triphosphate (G3P) is oxidized with G3P oxidase specific to G3P, and the hydrogen peroxide (H ₂ O ₂ ) generated at that time is oxidized with peroxidase and redox coloring reagent TOOS (N-ethyl- It can be quantified using N- (2-hydroxy-3-sulfopropyl) -3-methoxyaniline sodium) reagent (available from Dojindo Laboratories, Inc.). Furthermore, in order to increase the sensitivity, an enzyme cycle method is used in which the dihydroxyacetone phosphate generated by the oxidation reaction of G3P is returned to G3P again with a reductase, and again oxidized with G3P oxidase to increase the amount of H ₂ O ₂ generated. (Kishimoto T. et al., Clin Chim Acta. 2003 Jul 1; 333 (1): 59-67). In this method, an automatic analyzer can be used, and measurement can be performed in the same manner as a normal clinical test.

  The enzyme measurement method has an advantage that it can be carried out more easily at the clinical site. This method can be detected if the lysophospholipid in the sample is 0.03 μM or more.

  The mass spectrometry can be performed according to the method described in, for example, M. Okudaira et al., J. Lipid Res., 2014, 55: 2178-2192.

  Mass spectrometry can detect each molecular species, has high detection sensitivity, can detect a very small amount of molecular species, and can detect a large number of specimens in parallel.

  Analytical instruments that can be used in mass spectrometry are not particularly limited. For example, when TSQ Quantum Ultra (Thermo Fisher Scientific) is used as a mass spectrometer, each molecular species of lysophospholipid LPA, LPC, and LPI in a sample is used. Can be detected if each is 10 nM or more. In addition, when TSQ Quantiva (Thermo Fisher Scientific) is used as a mass spectrometer, it can be detected if each molecular species of lysophospholipid LPA, LPC, and LPI in the sample is 0.5 nM or more.

  Therefore, which detection method is used can be appropriately determined according to the number of specimens, the type of molecular species to be measured, and the like.

  The method of the present invention can further comprise comparing the detection result of lysophospholipid in the sample with a reference value for determining the presence and / or severity of neuropathic pain. The “reference value” is obtained as a standard value or range in advance as a boundary value for determining the presence and / or severity of pain, or a control sample without pain, or a value in the case of mild pain. Can do. The reference value can be set, for example, for each molecular species of total LPA, total LPC, LPA and LPC. For example, in the patient group divided into two groups according to the presence or absence of pain or severity, detection results for individual molecular types of total LPA, total LPC, LPA and LPC are obtained, and the statistically obtained cutoff value is used as a reference Can be used as a value.

  Based on the detection result from the patient-derived sample, it is possible to determine the presence or absence of pain or the severity of pain using the reference value. For example, if a value higher than the reference value for determining whether or not neuropathic pain is present, the patient can be determined to have pain symptoms, and the severity of neuropathic pain can be determined. If a value higher than the reference value for is found, the patient can be determined to have severe pain.

  In the method of the present invention, the detection result as described above is further converted into a pain assessment by a neuropathic pain assessment questionnaire (NPSI) and / or a Zurich lameness questionnaire (ZCQ), and a morphological severity by nuclear magnetic resonance imaging. Data for objective evaluation can be obtained in combination with evaluation and / or nerve damage evaluation in which phosphorylated neurophilament heavy chain (pNFH) is determined as a marker.

  NPSI is a method for evaluating the subjective severity of pain intensity, and ZCQ is a method for evaluating the degree of pain aversion caused by walking for intermittent claudication. Both methods are commonly used to quantify the subjective assessment of pain.

  As a result of examining the correlation between these evaluation methods and the method of the present invention, LPA and LPC generally have high values in the severe group in the NPSI evaluation and ZCQ evaluation, especially in the severe group patient in NPSI, LPA_18_2, LPC_14_0, LPC_16_0, LPC_16_1, LPC_18_0, LPC_18_1, LPC_18_2, LPC_20_4, LPC_20_5, and LPC_22_6 showed significantly higher lysophospholipid levels than the mild group. In addition, the patients in the severe group with ZCQ were significantly higher in the total LPA, LPA_18_1, LPA_18_2, LPA_20_4, LPC_16_0, LPC_16_1, LPC_18_0, LPC_18_1, LPC_18_2, LPC_20_4, LPC_20_5, and LPC_22_6, compared with the slightly higher lipid level. It was observed.

  A method reported by Schizas et al. (C. Schizas et al., SPINE, Vol.35, No.21, pp 1919-1924, is known) for evaluating the morphological severity of lumbar spinal canal stenosis. 2010). In this method, the degree of nerve compression is classified into grades A to D based on images obtained using nuclear magnetic resonance imaging (MRI).

  When the correlation between the above morphological severity assessment and the method of the present invention was examined, LPA and LPC generally showed a high value in the severe group in the NPSI assessment and ZCQ assessment, in particular, total LPA, LPA_16_0 , LPA_18_0, LPA_18_1, LPA_20_4, LPC_14_0, LPC_16_1, LPC_18_0, LPC_18_2, and LPC_22_6 showed significantly higher lysophospholipid levels in patients in the severe group with morphological grading compared to the mild group.

  On the other hand, phosphorylated neurophilament heavy chain (pNFH), a protein molecule contained in the myelin sheath of the central nervous system, is released from the axon in a phosphorylated state during axonal injury, resulting in cerebrospinal fluid (CSF) ) And leakage into the blood. Therefore, the presence of pNHF in cerebrospinal fluid (CSF) and blood is considered to be an indicator of nerve damage, and kits for detecting and quantifying pNFH in biological samples are available, for example, Funakoshi Co., Ltd., Cosmo Bio Co., Ltd. Commercially available from companies. However, the present inventors have found that the method of the present invention is more effective than the evaluation of nerve damage using pNFH as an index in order to objectively evaluate patient pain.

  That is, both the Schizas method and the pNFH method can indicate the degree of nerve damage, but these may not necessarily correlate with the presence and / or severity of pain as a patient's subjective symptoms. obtain.

  Therefore, the method of the present invention can be combined with the above-described existing evaluation method to provide data for diagnosis by combining the presence / absence and severity of neuropathy with the presence / absence and severity of pain, and lead to appropriate treatment. Make it possible.

  The present invention also provides a diagnostic marker for the presence and / or severity of neuropathic pain that includes an increase in the amount of LPA and / or LPC in the cerebrospinal fluid as an indicator.

  As described above, the findings of the present inventors revealed that an increase in the amount of LPA and / or LPC in cerebrospinal fluid is closely correlated with the presence and / or severity of neuropathic pain in the patient. did. Therefore, the total LPA, or LPA_16_0, LPA_18_0, LPA_18_1, LPA_18_2, LPA_20_4, and LPA_22_6, and LPC_14_0, LPC_16_0, LPC_16_1, LPC_18_0, LPC_18_1, LPC_18_2, LPC_20_4, LPC_20_4, and LPC_20_5 It can be used as a diagnostic marker for the determination of the presence and / or severity of neuropathic pain.

  The present invention also includes a reagent for quantitative detection of LPA and / or LPC in cerebrospinal fluid, reference value information in a control sample or control sample, and determination of the presence and / or severity of neuropathic pain. Providing a kit for

  The kit of the present invention can be used for carrying out the above-described method of the present invention. The “reagent” for quantitative detection of LPA and / or LPC is not particularly limited, but for example, reagents used in the above enzyme method, specifically, lysophospholipase, G3P oxidase, peroxidase, reductase And redox coloring reagents such as TOOS (N-ethyl-N- (2-hydroxy-3-sulfopropyl) -3-methoxyaniline sodium) reagent. Moreover, reagents, such as a buffering agent and a pH adjuster, can also be included as needed. The kit can also include a control sample containing a predetermined amount of LPA and / or LPC as a comparative control that can be measured simultaneously. The control sample need not be derived from cerebrospinal fluid and may be artificially prepared. The detected value of LPA and / or LPC in the control sample may be within the above reference value or range. Alternatively, LPA and / or LPC detection values (reference values) information that may be possessed by a control sample, such as a sample from a patient without neuropathic pain or a patient with mild pain, such as an enzymatic method or Detection results assumed in mass spectrometry can be included in the kit as information, for example, in the form of instructions for use.

  The method of the present invention also enables an objective assessment of pain due to interstitial cystitis. “Pain caused by interstitial cystitis” is bladder pain that the patient feels in the lower abdomen. This pain is similar to pain caused by infectious cystitis or bladder cancer. However, interstitial cystitis is difficult to diagnose because it lacks abnormal urinary findings seen in other diseases and requires experience in determining endoscopic findings. The method of the present invention can objectively evaluate “pain caused by interstitial cystitis” and discriminate it from other diseases by detecting the concentration of LPA and / or LPC in cerebrospinal fluid.

  In one embodiment, the LPA to be detected is one or more selected from LPA_16_0, LPA_18_1, and LPA_18_2, that is, one, two, or three. Although it does not limit, it is suitable to detect at least LPA_16_0 and / or LPA_18_2 among the above-mentioned molecular species.

  In one embodiment, the LPC to be detected is one or more selected from LPC_16_0, LPC_16_1, LPC_18_0, LPC_18_1, LPC_18_2, and LPC_22_6, that is, one, two, three, four, five or six. is there. Although not limited, it is preferable to detect at least LPC_16_0, LPC_18_1, and / or LPC_22_6 among the above molecular species.

  The method of the present invention is effective for interstitial cystitis using LPA present at a relatively low concentration in cerebrospinal fluid and / or LPC present at a relatively high concentration as a marker compared to other diseases. Objective pain is determined. The molecular species to be detected may be one or more of the above, but by combining a plurality of molecular species, the reliability (sensitivity and specificity) of the detection result can be further improved. Detection and quantification of LPA and LPC in a sample can be performed by, for example, but not limited to, mass spectrometry, particularly LC-MS / MS combining liquid chromatography and tandem mass spectrometry.

  The method of the present invention can further comprise comparing the detection result of lysophospholipid in the sample with a reference value for determining pain caused by interstitial cystitis. A boundary value for determining the presence or absence of pain, or a control sample without pain, or a value in the case of pain caused by another disease such as infectious cystitis, bladder cancer, etc., can be acquired in advance as a reference value it can. The reference value can be set, for example, for each molecular species of total LPA, total LPC, LPA and LPC. For example, in the patient group divided into two groups depending on the presence or absence of pain, detection results for individual molecular species of total LPA, total LPC, LPA and LPC are obtained, and the cut-off value obtained statistically is used as a reference value be able to.

  Based on the detection result from the patient-derived sample, the presence or absence of the patient's pain can be determined using the reference value. For example, total LPA, or each molecular species of LPA, for example, one or more molecular species selected from LPA_16_0, LPA_18_1, and LPA_18_2, a value lower than a reference value for determining pain caused by interstitial cystitis Can be determined to be interstitial cystitis. Alternatively, the determination of pain due to interstitial cystitis in one or more molecular species selected from total LPC, or each molecular species of LPC, for example, LPC_16_0, LPC_16_1, LPC_18_0, LPC_18_1, LPC_18_2, and LPC_22_6 If the value is higher than the reference value for the patient, it can be determined that the patient has interstitial cystitis. The determination can be made only by reducing the amount of LPA compared to the reference value, or can be made only by increasing the amount of LPC compared to the reference value. Furthermore, a decrease in the amount of LPA compared to the reference value and an increase in the amount of LPC can be detected and determined simultaneously.

The present invention also provides a diagnostic marker for pain caused by interstitial cystitis, which includes, as an indicator, a reduction in the amount of LPA and / or an increase in the amount of LPC in cerebrospinal fluid.
As described above, it has been found that reducing the amount of LPA and / or increasing the amount of LPC in cerebrospinal fluid correlates closely with pain due to interstitial cystitis in the patient. Accordingly, a reduction in the amount of one or more LPAs selected from LPA_16_0, LPA_18_1, and LPA_18_2 and / or the amount of one or more LPCs selected from LPC_16_0, LPC_16_1, LPC_18_0, LPC_18_1, LPC_18_2, and LPC_22_6 Augmentation can be used as a diagnostic marker for the determination of pain due to interstitial cystitis.

  The present invention also provides a reagent for quantitative detection of LPA and / or LPC in cerebrospinal fluid, reference value information in a control sample or a control sample, and determination of pain caused by interstitial cystitis. Provide kit.

  Hereinafter, the present invention will be described in more detail with reference to examples, but the present invention is not limited to these examples. All statistical analyzes were performed using JMP 12 (SAS Institute, Cary, NC, USA), and analysis of variance was conducted to examine the correlation between CSF levels of lysophospholipids and the severity of lumbar spinal stenosis. Later, the Tukey Honest Significant Differences test was used.

[Example 1] Evaluation of the presence or absence of neuropathic pain 53 patients with subjective symptoms of pain due to lumbar spinal canal stenosis (LSS) (hereinafter referred to as "patient group" for convenience) and no subjective symptoms The test was conducted on 12 patients (hereinafter referred to as “control group” for convenience).

  Cerebrospinal fluid (CSF) collected from each patient at the time of myelography was used as a sample and the enzyme according to the method described in Kishimoto T. et al., Clin Chim Acta. 2003 Jul 1; 333 (1): 59-67. The total LPA and total LPC amounts in the sample were quantified by the measurement method.

  In addition, using CSF 1 ml as a sample, according to the method described in M. Okudaira et al., J. Lipid Res., 2014, 55: 2178-2192, lysophospholipid LPA, LPC, LPI and sphingosine monophosphate (S1P) Quantification was performed.

  Specifically, the CSF sample was mixed with 10 times the amount of methanol and an internal standard, and sonicated. After centrifugation at 21,500 × g, the supernatant was recovered and used for LC-MS analysis. Next, 20 μL of the methanol extract was separated by Nanospace LC (Shiseido) using a C18 CAPCELL PAK ACR column (1.5 × 250 mm, Shiseido) (solvent A: 5 mM ammonium formate aqueous solution; solvent B: 95% [v / v] acetonitrile. Medium 5 mM ammonium formate).

  Eluent is ionized using an ESI probe and parent and fragment ions in positive ion mode using the TSQ Quantum Ultra Triple Quadrupole Mass Spectrometer (Thermo Fisher Scientific) or TSQ Quantiva Triple Quadrupole Mass Spectrometer (Thermo Fisher Scientific) Monitored. Table 1 shows m / z used for each lysophospholipid molecular species. In this example, since lipid was extracted under neutral conditions, 1-acyl-2-lysophospholipid and 2-acyl-1-lysophospholipid were not separated.

  For each lysophospholipid molecular species, 12 acyl chains (14_0, 16_0, 16_1, 18_0, 18_1, 18_2, 18_3, 20_3, 20_4, 20_5, 22_5 and 22_6) were monitored. As a result, 5 LPA (16_0, 18_0, 18_1, 18_2, 20_4), 8 LPC (14_0, 16_0, 16_1, 18_0, 18_1, 18_2, 20_4, 22_6) were obtained from CSF from patients and controls. And one S1P (C18) was detected.

  As a result of comparing the total LPA and total LPC in the sample and the detected values of each of the above molecular species between the patient group and the control group, LPA and LPC generally showed high values in the patient group. The results are shown in FIGS.

  Subsequently, in order to determine the significance of the obtained results, the Mann-Whitney U test and the Wilcoxon rank sum test were simultaneously performed. Table 2 below shows the U value, W value, Z value and asymptotic significance obtained for each molecular species.

  As can be seen from the results in Table 2, all of the eight types of LPA, 16_0 and 18_1, and LPC, 14_0, 16_0, 16_1, 18_0, 18_1, 18_2, 20_4, and 22_6 are statistically significantly higher in the patient group. Detected in CSF. On the other hand, sphingosine monophosphate (S1P_C18) did not give significant results. Among them, LPA16_0, LPA18_1, LPC20_4, and LPC_22_6 did not overlap the 95th percentile range with the control group, and were considered more useful as diagnostic indicators.

  Subsequently, ROC curves were prepared for the total LPA and each of the above molecular species, cut-off values were set, and the area under the ROC curve, sensitivity, and specificity were calculated. The results are shown in Table 3.

  From the results of Table 3, it was revealed that the detection results of the total LPA and the detected molecular species of LPA and LPC can be markers that can objectively determine the presence or absence of pain, respectively. In particular, with LPA, high sensitivity and specificity were obtained with total LPA and ROC area of about 0.8 or more in two types of 16_0 and 18_1. On the other hand, in LPC, all 8 types of 14_0, 16_0, 16_1, 18_0, 18_1, 18_2, 20_4, and 22_6 obtained an ROC area of about 0.9 or more, and very high sensitivity and specificity. It should be noted that good results were not obtained with sphingosine monophosphate.

Of the above molecular species, LPA_16_0 and LPA_18_1 were selected, and the following discriminant was created for the presence of neuropathic pain.
Y = -1.139 + [21.449 * (LPA16_0)] + [0.455 * (LPA18_1)]
In the above formula, when Y <0, it can be determined as neuropathic pain. In this case, the sensitivity is 54.7% and the specificity is 84.2%.

Moreover, LPC_20_4 and LPC_22_6 were selected from the above molecular species, and the following discriminant was created for the presence of neuropathic pain.
Y '=-1.133+ [7247.454 * (LPC20_4)] + [-2814.877 * (LPC22_6)]
In the above formula, when Y ′ <0, it can be determined that it is neuropathic pain. In this case, the sensitivity is 83.0% and the specificity is 100%.

In addition, among the above molecular species, LPA_16_0, LPA_18_1, LPC_20_4, and LPC_22_6 were selected, and the following discriminant was created for the presence of neuropathic pain.
Y '' =-1.214+ [10.743 * (LPA16_0)] + [-48.783 * (LPA18_1)] + [9486.401 * (LPC20_4)]
+ [-2402.981 * (LPC22_6)]
In the above formula, when Y ″ <0, it can be determined that the pain is neuropathic. In this case, the sensitivity is 90.5% and the specificity is 94.7%.

  In addition, 10 types of lysophospholipids (LPA_16_0, LPA_18_1 and LPC_14_0, LPC_16_0, LPC_16_1, LPC_18_0, LPC_18_1, LPC_18_2, LPC_20_4, LPC_22_6) were selected, and as a result, nerve was detected with a sensitivity of 100% and specificity of 94.7%. The presence or absence of sexual pain could be determined. That is, it was shown that sensitivity and specificity can be improved by determining the detection results of a plurality of molecular species in combination.

  On the other hand, the concentration of phosphorylated neurofilament heavy chain (pNF-H) in cerebrospinal fluid from the same patient group and control group was measured and compared between the patient group and the control group. The cut-off value was 380.5 pg / ml. In this case, the area under the ROC curve was 0.78, the sensitivity was 0.70, and the specificity was 0.929, indicating that the method of the present invention can be a better index.

[Example 2] Evaluation of severity of neuropathic pain 1-Correlation with NPSI evaluation 28 patients (15 males and 13 females) with subjective symptoms of pain due to lumbar spinal stenosis (LSS) 49-81 years old, average age 70.9 years).

  The lysophospholipid was quantified in the same manner as in Example 1, and total LPA and LPC were quantified by an enzyme measurement method. At the same time, 12 acyl chains (14_0, 16_0, 16_1, 18_0, 18_1, 18_2, 18_3, 20_3, 20_4, 20_5, 22_5 and 22_6) were monitored by mass spectrometry for each lysophospholipid molecular species. As a result, the CSF from the subjects was 6 LPA (16_0, 18_0, 18_1, 18_2, 20_4, 22_6), 1 LPI (18_0), 9 LPCs (14_0, 16_0, 16_1, 18_0, 18_1, 18_2, 20_4, 20_5, 22_6) and one kind of S1P (C18) were detected.

  On the other hand, for all patients, the Neuropathic Pain Assessment Questionnaire (NPSI) was used to assess the clinical severity of neuropathic pain due to lumbar spinal canal stenosis, and patients were identified with their severity of symptoms. Classified into 2 groups according to pain grading. Patients with a score of 21 or less (n = 14) were classified as mild (mild), and patients with a score of 22 or more (n = 14) were defined as severe (severe).

  As a result of evaluating the amount of lysophospholipid in the cerebrospinal fluid samples of the subjects together with the clinical severity evaluated by NPSI, LPA and LPC were generally high in the severe group. The results of plotting the concentration of lysophospholipid in the cerebrospinal fluid from each subject in the mild group and severe group for each molecular species are shown in FIG. 3 and FIG.

  In particular, in 1 type of LPA (LPA_18_2) and 9 types of LPC (LPC_14_0, LPC_16_0, LPC_16_1, LPC_18_0, LPC_18_1, LPC_18_2, LPC_20_4, LPC_20_5, LPC_22_6) compared with severe group patients with NPSI, Significantly higher lysophospholipid levels were seen.

  ROC curves were prepared for total LPA and each molecular species, cut-off values were set, and the area under the ROC curve, sensitivity, and specificity were calculated. The results are shown in Table 4.

  From the results of Table 4, it became clear that the detection results of the total LPA and the detected LPA and LPC molecular species can be markers that can objectively determine the severity of pain, respectively. . It should be noted that good results were not obtained with sphingosine monophosphate.

  From the results in Table 4, select 12 types of lysophospholipids (LPA_18_2, LPA_20_4, LPA_22_6 and LPC_14_0, LPC_16_0, LPC_16_1, LPC_18_0, LPC_18_1, LPC_18_2, LPC_20_4, LPC_20_6, LPC_20_6, LPC_20_6, LPC_20_5, PC) As a result of discriminant analysis, it was possible to determine the severity of neuropathic pain with a sensitivity of 100% and a specificity of 90% or more. That is, it was shown that sensitivity and specificity can be improved by determining the detection results of a plurality of molecular species in combination.

  In addition, considering the coefficient of discriminant, it is suggested that LPC_18_0, LPA_18_2, and LPA_22_6 are the most important, and then LPC_16_0 and LPC_22_6 are important in order to further improve the correlation with NPSI evaluation. .

[Example 3] Evaluation of severity of neuropathic pain 2-correlation with ZCQ evaluation The detection result of lysophospholipid in the sample obtained in Example 2 is evaluated by the Zurich lameness questionnaire (ZCQ) It was evaluated in conjunction with the clinical severity of pain-related lower limb movement disorders.

  For all patients, ZCQ was used and patients were classified into two groups according to their symptom severity and pain grading. Patients with a score of 20 or less (n = 14) were classified as a mild group, and patients with a score of 21 or more (n = 14) were defined as a severe group.

  As a result, the patients in the severe group with ZCQ showed high values in the severe group as a whole with LPA and LPC. The results of plotting the concentration of lysophospholipid in the cerebrospinal fluid from each subject in the mild group and severe group for each molecular species are shown in FIGS.

  In particular, in severe LPC, 3 cases of LPA (LPA_18_1, LPA_18_2, LPA_20_4) and 8 types of LPC (LPC_16_0, LPC_16_1, LPC_18_0, LPC_18_1, LPC_18_2, LPC_20_4, LPC_20_5, LPC_22_6), severe in ZCQ patients, Significantly higher lysophospholipid levels were seen compared to the group.

  ROC curves were prepared for total LPA and each molecular species, cut-off values were set, and the area under the ROC curve, sensitivity, and specificity were calculated. The results are shown in Table 5.

  From the results of Table 5, it became clear that the detection results of the total LPA and the detected molecular species of LPA and LPC can be markers that can objectively determine the severity of pain, respectively. . It should be noted that good results were not obtained with sphingosine monophosphate.

  From the results in Table 5, 13 types of lysophospholipids (LPA_18_1, LPA_18_2, LPA_20_4, LPA_22_6 and LPC_14_0, LPC_16_0, LPC_16_1, LPC_18_0, LPC_18_1, LPC_18_2, LPC_20_4, LPC_20_4, LPC_20_5, LPC_20_4, LPC_20_4, LPC_20_4 As a result of discriminant analysis, it was possible to determine the severity of neuropathic pain with a sensitivity of 100% and a specificity of 90% or more. That is, it was shown that sensitivity and specificity can be improved by determining the detection results of a plurality of molecular species in combination.

  In addition, considering the coefficient of discriminant, it is suggested that LPC_16_0, LPC_18_0, LPC_22_6 is the most important, and then LPA_18_1, LPC_18_1, LPA_20_4 is important in order to further improve the correlation with ZCQ evaluation. It was done.

[Example 4] Evaluation of severity of neuropathic pain 3-Correlation with morphological severity The detection result of lysophospholipid in the sample obtained in Example 2 was measured using nuclear magnetic resonance imaging. It was evaluated in conjunction with the morphological severity of lumbar spinal canal stenosis evaluated.

  For all patients, nuclear magnetic resonance imaging was used to assess the morphological severity of lumbar spinal canal stenosis by the method of Schizas et al. And according to its morphological grading (CSF / root ratio) Classified into groups. Grades A and B were mild (n = 10) and Grades C and D were severe (n = 18).

  As a result, morphological grading of patients in the severe group, LPA and LPC were generally high in the severe group. The results of plotting the concentration of lysophospholipid in the cerebrospinal fluid from each subject in the mild group and severe group for each molecular species are shown in FIGS.

  Especially in total LPA, 4 types of LPA (LPA_16_0, LPA_18_0, LPA_18_1, LPA_20_4) and 5 types of LPC (LPC_14_0, LPC_16_1, LPC_18_0, LPC_18_2, LPC_22_6) in patients with severe morphological grading, mild Significantly higher lysophospholipid levels were seen compared to the group.

  ROC curves were prepared for each of the above molecular species, cut-off values were set, and the area under the ROC curve, sensitivity, and specificity were calculated. The results are shown in Table 6.

  From the results of Table 6, it became clear that the detection results of the total LPA and the detected molecular species of LPA and LPC can be markers that can objectively determine the severity of pain, respectively. . It should be noted that good results were not obtained with sphingosine monophosphate.

  From the results of Table 6, 12 lysophospholipids (LPA_16_0, LPA_18_0, LPA_18_1, LPA_20_4, and LPC_14_0, LPC_16_0, LPC_16_1, LPC_18_0, LPC_18_1, LPC_18_2, LPC_20_4, LPC_20_4, LPC_20_4, LPC_20_4, LPC_20_4, LPC_20_4 As a result of discriminant analysis, it was possible to determine the severity of neuropathic pain with a sensitivity of 100% and a specificity of 90% or more. That is, it was shown that sensitivity and specificity can be improved by determining the detection results of a plurality of molecular species in combination.

  Considering the discriminant coefficients, it is most important to include LPC_18_1 and LPA_20_4, followed by LPC_16_1, LPA_16_0, and LPC_16_0 in order to further enhance the correlation with morphological evaluation. It was done.

[Example 5] Evaluation of pain due to interstitial cystitis Brains from 13 patients with interstitial cystitis having subjective symptoms of pain in the lower abdomen and 22 patients with bladder cancer and urolithiasis without pain Spinal fluid (CSF) was collected, and lysophospholipid LPA and LPC molecular species were quantified in the same manner as in Example 1. Table 7 shows the results of comparison with the bladder cancer patient group regarding the cerebrospinal fluid concentration of each molecular species in the sample of the interstitial cystitis patient group. In the table, → indicates that the cerebrospinal fluid concentration is almost the same, ↑ indicates that the concentration is high in patients with arthritic cystitis, and ↑ indicates that the concentration is low in patients with arthritic cystitis.

  From the results of Table 7, it was shown that in the cerebrospinal fluid of patients with interstitial cystitis, the LPA concentration tends to be lower and the LPC concentration tends to be higher than those of bladder cancer patients. Specifically, in LPA, the concentrations of LPA_16_0, LPA_18_1, and LPA_18_2 were low, and the concentrations of LPC_16_0, LPC_16_1, LPC_18_0, LPC_18_1, LPC_18_2, and LPC_22_6 were high. In particular, among the above molecular species, the results of low concentrations of LPA_16_0 and LPA_18_2 and the results of high concentrations of LPC_16_0, LPC_18_1 and LPC_22_6 were significant. Note that there was no difference in the concentration of autotaxin (ATX), an enzyme that hydrolyzes LPC to produce LPA.

  Chronic neuropathic pain is recognized as a disease with high socioeconomic costs including medical expenses. The method of the present invention can objectively quantify neuropathic pain that has not been clearly clarified so far, and can be linked to more appropriate treatment thereafter, and is very useful. . Furthermore, according to the method of the present invention, the pain caused by interstitial cystitis can be objectively evaluated, leading to earlier diagnosis and more accurate treatment.

Claims (21)

  A method for acquiring data for objective evaluation of pain, comprising detecting lysophosphatidic acid (LPA) and / or lysophosphatidylcholine (LPC) in a sample derived from human cerebrospinal fluid.   The method according to claim 1, wherein the detection of LPA and / or LPC is detection for each molecular species.   The method according to claim 1 or 2, wherein the pain is neuropathic pain, and the objective evaluation is a determination of the presence and / or severity of neuropathic pain.   The method according to any one of claims 1 to 3, wherein the LPA is at least one selected from total LPA, LPA_16_0, LPA_18_0, LPA_18_1, LPA_18_2, LPA_20_4, and LPA_22_6.   The method according to claim 4, wherein at least LPA_16_0 and / or LPA_18_1 is detected.   The method according to any one of claims 1 to 5, wherein the LPC is at least one selected from LPC_14_0, LPC_16_0, LPC_16_1, LPC_18_0, LPC_18_1, LPC_18_2, LPC_20_4, LPC_20_5, and LPC_22_6.   The method according to claim 6, wherein at least LPC_20_4 and / or LPC_22_6 is detected.   The method according to any one of claims 1 to 7, wherein at least LPA_16_0, LPA_18_1, LPC_20_4, and LPC_22_6 are detected.   The method according to any one of claims 1 to 8, which is detected by an enzymatic method or mass spectrometry.   The method according to any one of claims 1 to 9, further comprising comparing the detection result with a reference value for determining the presence and / or severity of neuropathic pain.   Based on the results of the evaluation, the neuropathic pain assessment questionnaire (Neuropathic Pain Symptom Inventory, NPSI) and / or the Zurich Claudication Questionnaire (ZCQ), pain assessment, morphological severity assessment by nuclear magnetic resonance imaging, And / or data for objective evaluation in combination with nerve damage evaluation in which phosphorylated neurophilament heavy chain (pNFH) is determined as a marker. Method.   A diagnostic marker for determining the presence and / or severity of neuropathic pain, which includes an increase in the amount of LPA and / or LPC in cerebrospinal fluid as an indicator.   A kit for determining the presence and / or severity of neuropathic pain, comprising a reagent for quantitative detection of LPA and / or LPC in cerebrospinal fluid, reference value information in a control sample or a control sample.   The method according to claim 1 or 2, wherein the pain is pain caused by interstitial cystitis.   The method according to claim 14, wherein the LPA is one or more selected from LPA_16_0, LPA_18_1, and LPA_18_2.   The method according to claim 15, wherein at least LPA_16_0 and / or LPA_18_2 is detected.   The method according to any one of claims 14 to 16, wherein the LPC is at least one selected from LPC_16_0, LPC_16_1, LPC_18_0, LPC_18_1, LPC_18_2, and LPC_22_6.   The method according to claim 17, wherein at least LPC_16_0, LPC_18_1 and / or LPC_22_6 is detected.   The method according to any one of claims 14 to 18, further comprising comparing the detection result with a reference value for determining pain due to interstitial cystitis.   A diagnostic marker for the determination of pain caused by interstitial cystitis, which includes, as an index, a reduction in the amount of LPA and / or an increase in the amount of LPC in cerebrospinal fluid.   A kit for the determination of pain due to interstitial cystitis, comprising a reagent for quantitative detection of LPA and / or LPC in cerebrospinal fluid, reference value information in a control sample or a control sample.

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