CN101150983B - Systems and methods using nuclear magnetic resonance (NMR) spectroscopy to evaluate pain and degenerative properties of tissue - Google Patents

Abstract

NMR spectroscopy is performed on intervertebral disc tissue. Extent of degeneration is determined based on the NMR spectroscopy. Correlation between NMR spectral regions and pain are made. Accordingly, NMR spectroscopy is used to determine location and/or extent of at least one of disc degeneration, or discogenic pain.

Description

Nuclear magnetic resonance, NMR (NMR) is utilized to compose the system and method for assessment tissue irritation and degenerative character

About the cross reference of application

This application claims the U.S.Provisional Serial 60/648 submitted on January 28th, 2005, the priority of 241, by its overall introducing the present invention as a reference, and require the U.S.Provisional Serial 60/731 submitted on November 15th, 2005, the priority of 110, also by its overall introducing the present invention as a reference.

About the statement of federal sponsored research or exploitation

The present invention carries out under governmental support, and it is R01-AG17762 and R21-AR51048 that National Institutes of Health subsidizes license number.Government has specific rights of the present invention.

The reference content that the data that CD is submitted comprises

Inapplicable

Material declaration protected by copyright

A part of material of patent document is by the copyright protection of the U.S. with other National copyright laws.When the file that U.S.Patent & Trademark Office discloses or record, copyright owner not reproduction by anyone's facsimile copy patent document or patent discloses, in any case but but will retain whole copyright in other.Copyright owner can not abandon its any right accordingly to maintain the confidentiality of this patent documentation, comprises and does not limit its right according to the regulation of the 1.14th article, CFR the 37th chapter.

Technical field

The invention belongs to the imaging technique of the tissue relevant with skeletal joint, particularly about the identification and sign with skeletal joint, pain or the two relevant medical condition.More particularly, the present invention relates to using NMR (NMR) spectrum to the chemistry organizing medical condition relevant, molecule, structure or other signal identify, locate and characterize, medical condition is such as dependent degeneration or the pain of skeletal joint (as spine) herein.

Background technology

The low back pain that lumbar vertebra is correlated with have impact on 60-80% just in the American common medical problem of ageing, and intervertebral disc degeneration (IVDD) is its main reason.Intervertebral disc is a kind of pliable and tough fibrocartilage structure, can bear pressure and be convenient to vertebra and move.Healthy intervertebral disc is made up of three kinds of particular organization's compositions: 1) fibrous ring, and circumferentially shape is closely filled in intervertebral disc periphery and makes it have flexible collagenous region; 2) vertebral pulp, is positioned at a kind of moisture Dan Baiduotang proteoglycan PG gel at intervertebral disc center, during pressurized can radially expanding support fibrous ring to keep its intensity and to prevent fibrous ring Bending Deformation under stress; 3) cartilage endplate, makes vertebral pulp separate with adjacent vertebrae.

Intervertebral disc degeneration with the physics of series of complex with the degradation process of chemistry for feature.The degree and the seriousness that describe IVDD clinically use Tang Pusen classification rule (Thompson Grading Scale) usually: after setup parameter, utilize X-ray radiography to observe intervertebral disc, by general morphology determination degree of degeneration.Have research group to report, along with the regression of intervertebral disc, its mechanical performance there occurs the transformation from " liquid state " behavior to " solid-state " behavior.The fixed charge density (FCD) of ambient water and biological chemical environment are also considered to have a great impact regression; Because the Dan Baiduotang proteoglycan PG of high electric charge can adsorbed water causing be organized swelling, directly affects compression resistance and the carrying spine ability of intervertebral disc.The difference of the general Lignum Rhamnellae level of soup (Thompson Grade) is reflected by the concentration change of each component (as collagen and Dan Baiduotang proteoglycan PG) of ring-shaped area and center.The degraded of biochemical degradation, the collagen stroma relevant with gene upregulation, the accumulative effect of machinery carrying are all considered to the degenerative processes that can cause intervertebral disc.

So contain a large amount of developments in science and technology and people's effort for many years to the identification of intervertebral disc degeneration with sign.But, one suitable, repeatably, the system and method for Noninvasive for characterizing pain, pain produces and intervertebral disc degeneration factor provides a useful medical tool.

It should be noted that equally in modem medical practices, pain is the phenomenon of a quite difficult diagnosis and location, particularly in arthralgia especially backache.Such as, but some target pain release therapy may be desirable, makes energy source point to and the nerve of Partial ablation pain.Determine it is a crucial main project with locating therapy position, often for want of necessary specificity and cannot accurately locating.Therefore, in all pain management, how successfully to make these treatment lock onto targets be challenges greatly.

Degenerative disc disease, although the one of the main reasons being the backache making people unable, is just badly in need of better can characterizing and locate the Method and kit for of medical condition to carry out a kind of disease condition example of suitable targeted therapy.Such as chronic back pain, may be caused by several potential basic reason.These reasons comprise as vertebral compression fracture, degenerative disc disease and intervertebral disk hernia.In addition, other arthralgia, such as vertebra and other skeletal joint are (as articulations digitorum manus, ankle joint, knee joint, hip joint, shoulder joint, carpal joint, elbow joint) also may because of multiple separately or the different potential cause that be combined with each other caused, be also difficult to their accurately location to carry out direct topical therapeutic.Any one or all sore places relevant to these joints may be positioned at connection or the buffer tissue (such as intervertebral disc) in joint, also may be positioned at bone, or be positioned at transitional region (such as the cartilage endplate of spinal cord and intervertebral disc intersection).

To the Noninvasive tool and method can determining and characterize the improvement that in-vivo tissue is degenerated, there is practical demand at present.Particularly with regard to joint part, the joint especially between vertebrae, a nearlyer step particularly spinal disc interior and around with regard to all the more so.

To the Noninvasive tool and method can determining and characterize the improvement of painful area in body, also there is practical demand at present.This particularly with regard to skeletal joint position, the joint especially between vertebrae, a nearlyer step particularly to spinal disc interior and around with regard to all the more so.

Summary of the invention

An aspect disclosed by the invention there is provided a kind of medical diagnostic system, it has the form of non-invasive imaging, be applicable to the chemical signal based on the correlative factor of a certain feature and the information that provides, this information can indicate the degree of this feature of tissue regions.

The collection of illustrative plates utilizing NMR spectral method to obtain and the collection of illustrative plates of annulus fibrosus disci intervertebralis and nuclear area sample closely similar.Along with the increase of the general Lignum Rhamnellae level of soup, from annulus fibrosus disci intervertebralis district and nuclear area sample collection of illustrative plates, can be observed significant change.Distinguished between the intervertebral disc that in collection of illustrative plates, the area ratio of N-acetyl group region and choline base region and the area ratio in choline base region and carbohydrate region make the general Lignum Rhamnellae level of soup increase, the overlap of single area ratio is minimum.

According to an embodiment of this aspect, this feature is relevant with pain.

According to another aspect of this embodiment, this feature is relevant with tissue deterioration.

According to another aspect of this embodiment, this system provides the useful information that can indicate osteoarthrosis correlated characteristic.

According to another aspect of this embodiment, this system provides the useful information that can indicate spinal joint correlated characteristic.

According to another aspect of this embodiment, this system provides the useful information that can indicate intervertebral disc correlated characteristic.

According to another embodiment, this system provides the useful information that can indicate disc degeneration degree.

According to another embodiment, this system provides the useful information of the pain position that patient can be indicated to stand.

According to another embodiment, this system at least in part based on tissue regions NMR spectrum and provide useful information.

Following another aspect of the present invention provides a kind of medical diagnostic system, its spectrum of nuclear magnetic resonance, NMR (NMR) based on the first area of tissue provides the useful information of this first area feature of instruction, and wherein this feature is relevant at least one in tissue deterioration and pain.

Following another aspect of the present invention provides a kind of medical diagnostic system, and its spectrum of nuclear magnetic resonance, NMR (NMR) based on the first area of tissue provides the useful information of the feature of this first area of instruction, and wherein this feature is relevant to the degeneration of tissue.

Following another aspect of the present invention provides a kind of medical diagnostic system, and its spectrum of nuclear magnetic resonance, NMR (NMR) based on the first area of tissue provides the useful information of the feature of this first area of instruction, and wherein this feature is relevant to pain.

Following another aspect of the present invention provides a kind of system of the feature for identifying or characterize skeletal joint linked groups first area.This system comprises a kind of processor, composes relevant data for processing in the mode of the useful information providing this tissue first area feature of instruction to organizing NMR.

Following another aspect of the present invention provides a kind of system that can identify or characterize the feature of the first area of skeletal joint tissue.This particular system comprises a kind of NMR spectra system, and for providing the NMR organizing first area to compose, and for providing the data relevant to collection of illustrative plates, this data format is suitable for being processed into and can indicates the useful information organizing first area feature.

Following another aspect of the present invention provides a kind of system of the feature for identifying or characterize skeletal joint linked groups first area.This system comprises a kind of essentially no invasive imaging modality, can in order to provide the imaging data relevant to organizing first region, and the data of this form are suitable for being processed into the useful information that can indicate tissue first provincial characteristics.

Following another aspect of the present invention provides a kind of system that can identify the feature of at least first area of intervertebral disc.This system comprises a NMR spectra system, and this system is suitable for providing composes relevant data to organizing first area NMR, and the form of these data is suitable for being processed into the useful information of the feature of the first area that can indicate tissue.

Following another aspect of the present invention provides a kind of for providing the system of useful information for diagnostic organization's first area feature, and this tissue is relevant to tissue deterioration or pain.This system comprises a computer-readable software program existed with computer-readable media form, can process and organize the nuclear magnetic resonance nmr of first area to compose the data obtained.This program can provide useful information further from the data of process.

Another aspect of the present invention provides a kind of system for identifying, characterizing or locate skeletal joint linked groups feature.This system can further include any one or the multiple various systems, pattern, embodiment, modification or the device that provide and their assembly herein.

A kind of pattern of system according to the invention, this system is suitable for the information providing instruction intervertebral disc feature at least partially.

Another aspect of the present invention provides a kind of system that can identify or characterize patient's osteoarthrosis linked groups feature.Which comprises at least following one of them: processor, relevant information can be composed with the mode process characterized to organizing NMR to be suitable for making the degree of this feature to be identified; NMR spectra system, can provide the relevant spectral information of tissue, and the degree of its feature can be identified and characterize; Or a kind of essentially no invasive imaging modality, can provide the information of tissue, make the degree of its feature be identified and to characterize.Or this system can comprise the combination of two or more above-mentioned parts.

An embodiment of system according to the invention, described information is relevant with the characteristic length at least partially of intervertebral disc.

Another aspect of the present invention provides a kind of system, for characterizing the characteristic length at least partially of intervertebral disc.Native system comprises one and is suitable for catching this part and is correlated with the NMR spectra system of spectral information.This wave spectrum can be provided for the information indicating this characteristic length at least in part.

According to another detailed description of the invention of aforesaid aspect and system model, each system is suitable for providing relevant information based on any one in annulus fibrosus disci intervertebralis part and nucleus pulposus portion or both.

According to another embodiment, this system can show the correlation curve of nuclear magnetic resoance spectrum, and a part for curve can provide information.

According to another embodiment, described information is suitable for the degree of degeneration distinguishing intervertebral disc.According to one preferred embodiment, described information is suitable for according to Tang Pusen grade distinguishing degree of degeneration.

According to another embodiment, described feature at least comprises one of following: pain, or at least relevant to pain factor.

According to another embodiment, described information is relevant to the ratio of N-acetyl group Q region resonance Q and choline Q region resonance Q in wave spectrum and the ratio of choline Q region resonance Q and carbohydrate Q region resonance Q.

According to another embodiment, described information and chondroitin sulfate or its metabolite or its catabolite relevant.

According to another embodiment, the T of the chemical constituent that described information is at least composed with intervertebral disc ₁and T ₂one in relaxation time relevant.

According to another embodiment, described feature at least comprises one of following: the degree of dehydration of intervertebral disc, the degree of decomposition of intervertebral disc proteoglycan matrix, and the degree of decomposition of collagen stroma.

According to another embodiment, described system also comprises the proton high resolution Magic angle spinning spectra system being suitable for generation information.

Another aspect of the present invention is a kind of method providing feature for identifying and characterize osteoarthrosis linked groups.Aforesaid one or more aspect, pattern, embodiment, modification, the device described therebetween or their combination, all can be used for improving this method.

The another kind of pattern of this aspect comprises the information of the degree of the feature at least partially providing instruction intervertebral disc.

Another aspect of the present invention is to provide a kind of method for identifying and characterize patient's osteoarthrosis linked groups, comprises at least one following steps: to be suitable for the relevant information allowing the mode process of identification or characteristic feature degree to organize NMR to compose; There is provided profile information from NMR spectra system, it is relevant to tissue, characteristic length is identified or characterizes; Or provide relevant organizational information from the basic imaging modality for Noninvasive, to make the degree of feature be identified or characterize.Or above-mentioned one or more step can combinationally use.

An embodiment of this aspect is included in the characteristic length at least partially described Information base being determined intervertebral disc.

Another aspect of the present invention is to provide a kind of method characterizing the characteristic length at least partially of intervertebral disc, and comprises and utilize NMR spectra system to catch the relevant wave spectrum of this part.

According to an embodiment and the pattern of aforementioned various distinct methods, produce described information based in the annulus fibrosus portion of intervertebral disc and nucleus pulposus portion or both.

In another embodiment, show the curve relevant to wave spectrum, wherein a part for curve provides information.

The basis that another embodiment is included in information is distinguished the degree of disc degeneration.One further embodiment be included in information basis on according to the degree of Tang Pusen grade distinguishing disc degeneration.

Another embodiment is included on the basis of information, is associated by intervertebral disc with the degree of pain, or is at least that a kind of factor relevant with pain is associated.

According to another embodiment, described information is relevant at least one in following ratio: the ratio of N-acetyl group Q region resonance Q and choline Q region resonance Q in collection of illustrative plates, the ratio of choline Q region resonance Q and carbohydrate Q region resonance Q.

According to another embodiment, described information and chondroitin sulfate or its metabolite or its catabolite relevant.

According to another embodiment, the T of described information and intervertebral disc chemical constituent wave spectrum ₁and T ₂at least one in relaxation time is relevant.

According to another embodiment, described feature at least with one of following about the degree of dehydration of intervertebral disc, the degree of decomposition of intervertebral disc proteoglycan matrix, and the degree of decomposition of collagen stroma.

Another embodiment comprises and utilizes proton high resolution Magic angle spinning spectroscopy system to produce information at least partly.

According to the further append mode of aforementioned various different aspect, be included on the basis of at least one in the chemical signal that the chemical signal that identifies that chemical signal, Dan Baiduotang proteoglycan PG that in tissue, lactate is correlated with are correlated with or collagen is correlated with, useful non-invasive analysis is carried out to the feature of tissue.Particularly preferred pattern comprises, and such as, compares the ratio between these signal characteristics identified, and the NMR resonance based on these factors in a preferred embodiment compares.The present invention further preferred embodiment in, carry out the comparison of these resonance two or more, such as, particularly compare their peak or other indicates the feature of its content degree, for providing the differentiation result of particular condition degree in indicating target tissue (as certain is degenerated or the degree of pain).

One, these aspects further preferred pattern comprise the application of NMR spectrum.In one embodiment, Wave Spectrum condition is included in more than 8Tesla (tesla) and operates instrument, and in another embodiment, magnetic field intensity is between 11 ~ 12Tesla.In another embodiment, instrument is about running between 4 ~ 10Tesla, particularly under about 7Tesla.In another embodiment, instrument is about running under such as about 3Tesla between 2 ~ 4Tesla.In another embodiment, instrument runs under below 3Tesla such as about 1.5Tesla.To the instrument of lower tesla, further embodiment relates to the signal utilizing wave filter increase, amplification or acquisition to drop in specific lock-in range, such as some specific resonant frequency peak that identification signal is locked, this signal is relevant with some specific factors for non-invasive diagnostic that will study.

Preferably each aspect aforementioned, pattern, embodiment, modification or device are used alone, and do not require to combinationally use with alternate manner.But the further combination of these modes and secondary combination are also all considered to be independent optimal way of the present invention.

The present invention further aspect is described in description part below, comprise the content do not limited in claim, detailed description is wherein to describe embodiments of the invention and preferred implementation, not limit the invention, although these are preferred embodiment described as providing value useful especially and use.

Accompanying drawing explanation

The present invention may be better understood with reference to the following drawings.Following each figure is only for illustrating.

The representational 1-D HR-MAS that Fig. 1 X, 1Y, 1Z show the general gloomy 1 grade of intervertebral disc (Fig. 1 X) of the soup obtained from annular region (collection of illustrative plates of Fig. 1 Y) and nuclear area (collection of illustrative plates of Fig. 1 Z) composes.

The representational 1-D HR-MAS that Fig. 2 X, 2Y, 2Z show the general gloomy 3 grades of intervertebral disc (Fig. 2 X) of the soup obtained from annular region (collection of illustrative plates of Fig. 2 Y) and nuclear area (collection of illustrative plates of Fig. 2 Z) composes.Distinguishable peak comprises: A: isoleucine, leucine, valine; B: lactate, isoleucine; C: alanine; D: isoleucine, leucine; E: lysine, leucine; F: the N-Acetyl resonance of chondroitin sulfate; G: glutamine; H: glutamic acid, proline; I: glutamine, hydroxyproline; J: lysine; K: gallbladder alkali; L: phosphocholine; M: hydroxyproline; N: glycine; O: the C-H resonance of chondroitin sulfate; P: gallbladder amine; The C-H resonance of parantheses Regional Representative chondroitin sulfate.

Fig. 3 X, the representational 1-D HR-MAS that 3Y, 3Z show the general gloomy 5 grades of intervertebral disc (Fig. 3 X) of the soup obtained from annular area (collection of illustrative plates of Fig. 3 Y) and nuclear area (collection of illustrative plates of Fig. 3 Z) composes.Distinguishable peak comprises: A: isoleucine, leucine, valine; B: lactate, isoleucine; C: alanine; D: isoleucine, leucine; E: lysine, leucine; F: the N-Acetyl resonance of chondroitin sulfate, proline, glutamic acid; G: glutamine; H: glutamic acid, proline; I: glutamine, hydroxyproline; J: lysine; K: gallbladder alkali; L: phosphocholine; M: hydroxyproline; N: glycine; O: the C-H resonance of chondroitin sulfate; P: cholamine.

As described in Table 1, the figure line of Fig. 4 A-4D represents the integral distribution of the N-acetyl group/choline (N-Acetyl/Cho) (Fig. 4 A) of fibrous ring and the N-acetyl group/choline (Fig. 4 C) of choline/carbohydrate (Cho/Carb) (Fig. 4 B) and vertebral pulp and choline/carbohydrate (Fig. 4 D) and the relation of the general Lignum Rhamnellae level of soup.Choline/carbohydrate shows maximum statistical significance.

The heat-insulating TOCSY that Fig. 5 A shows the rotor of healthy disc material synchronous composes, and incorporation time is 80ms.Trunnion axis is the superposition of projection, and vertical axis is high-resolution one dimension spectrum.The amino acid code of three letter representations is used for indicating amino acid whose intersection peak.

The heat-insulating TOCSY that Fig. 5 B shows the rotor of the intervertebral disc of degeneration synchronous composes, and incorporation time is 80ms.In the intervertebral disc collection of illustrative plates of degenerating, aminoacid and the signal containing the compound of choline all enhance, and do not strengthen in the collection of illustrative plates of the intervertebral disc of health.

Fig. 6 A illustrates the average spin lattice relaxation time of following compound: 0.9ppm: isoleucine; leucine; and valine; 1.32ppm: lactate; 1.49ppm: alanine; 2.04ppm: chondroitin sulfate N-acetyl group part, 3.21ppm: containing the compound of choline, 3.67ppm: the C-H on the carbohydrate residue be connected with chondroitin sulfate polymer.

Fig. 6 B illustrates the average spin spin relaxation time of above-claimed cpd.(*) represent because the signal intensity at peak is low cause having very large standard deviation.

Fig. 7 A, 7B show the mapping example obtained in the NMR experiment of particular vertebral disc.

Detailed description of the invention

Carry out more specific description below in conjunction with accompanying drawing, illustratively, implement instrument of the present invention, be included in the equipment that the content that shows from Fig. 1 X to Fig. 6 B and table 1 utilizes, Fig. 7 A-B and table 2 also reflect a kind of AD HOC of improvement.It should be noted that structure and the component detail of instrument have some differences, the particular step of method therefor and order also have some changes, but do not deviate from mutually with the basic conception announced herein.

The ability utilizing nuclear magnetic resoance spectrum to characterize disc degeneration in specific material or chemical composition is disclosed at this.Mark in nuclear magnetic resoance spectrum and the Noninvasive mutual relation between soup general Lignum Rhamnellae level obtained, this can extensively be applied in pharmacy and scientific research by model, produces huge effect.The present invention is special, and usefulness is, provides the identification of Noninvasive and characterizes the ability of the mark relevant with present position to disc degeneration state, and can also identify and characterize this mark and pain position or pain and produce particular kind of relationship between the factor.

With reference to experimentation and discussion described in one or more embodiments subsequently, the different aspect of the present invention, pattern, embodiment, modification and device clearerly can be understood.

Embodiment 1

1. summary

The object of this research is to determine that high-resolution evil spirit angle spin (HR-MAS) NMR composes the ability of the different phase distinguishing disc degeneration.Obtain 17 intervertebral disc from people's corpse, adopt a peacekeeping two dimension (TOCSY) ¹hHR-MAS spectrum is analyzed, and measures T ₁and T ₂relaxation time is to determine the change of chemical composition and the increase intervertebral disc chemical environment along with degradation level (the general Lignum Rhamnellae level of soup).Research great discovery comprise: the annular area of intervertebral disc and the collection of illustrative plates of nuclear area sample closely similar; Along with the increase of the general Lignum Rhamnellae level of soup, the annular area of intervertebral disc and the collection of illustrative plates of nuclear area sample there occurs visual significant change.The intervertebral disc that in collection of illustrative plates, the area ratio in N-acetyl base and choline district and the area ratio in acetyl base and carbohydrate district can make the general Lignum Rhamnellae level of soup increase is distinguished, and overlap between these single ratios is very little.The T of the chemical composition of intervertebral disc collection of illustrative plates ₁and T ₂relaxation time change seems to reflect the increase along with the general Lignum Rhamnellae level of soup, the change of the dehydration change of intervertebral disc and Dan Baiduotang proteoglycan PG, collagen stroma degree of decomposition.The result of this research proves, can utilize the chemical change that in body, spectral method detection occurs with intervertebral disc degeneration.

Carry out MRI (NMR (Nuclear Magnetic Resonance)-imaging) research in a few individuality herein, to characterize the feature of IVDD (intervertebral disc degeneration) better.T ₁and T ₂the MRI of weight is used to the structure analyzing intervertebral disc.Research proves, T ₂the increase that the decline of the signal intensity of weighting is degenerated with lumbar intervertebral disc.Along with the regression of sample, the T of water in degenerated cartilagenous ₁value can significantly reduce.Also observe the regression along with articular cartilage, the T of its water ₂relaxation time can change.By diffusion-weighted imaging research intervertebral disc and cartilage, find that the minimizing of water content is relevant with catagen phase.In the MRI of cervical vertebra, along with the increase at age, more serious concerning the situation that dehydration occurs all intervertebral disc.Have research group to infer, this caused not due to damage or recurrent stress, but owing to there occurs more balanced regression.The discovery of these NMR (Nuclear Magnetic Resonance)-imaging adds the accuracy of morphologic grading overall system, but by adopting the method based on the detectable chemical constituent of spectroscopic measurement in body, measurement degenerative states that can be more effective and quantitative.

Be starved of now a kind of technology of Noninvasive to characterize complete disc tissue metabolite component in better terrain.The traditional method measuring chemical composition needs to extract protein by biochemical method, and this understands disorganize and hampers further research (such as, biological assay or mechanical test).HR-MAS (magic angle spinning) nuclear magnetic resoance spectrum is a kind of technology of Noninvasive, has been used successfully to and has characterized various complete biological tissue's composition.By degraded by collagenase bovine nasal cartilage, we simulate cartilage degradation, and catabolite is studied by magic angle spinning nuclear magnetic resoance spectrum.This makes the Natural Degradation product of the similar cattle tissue of the amino acid product of collagen helix, which provides the regression model of people's tissue.But the differentiation level of the biochemistry relevant with the change of intermedium degree of degeneration, mechanical degeneration still general Lignum Rhamnellae level can characterize with single soup, and this method also highlighting needs ratio present more has illustrative staging hierarchy.

The object of this research is to show that HR-MAS spectrum can be used for evaluating the chemical change relevant with intervertebral disc degeneration.When carrying out In vivo NMR experiment to characterize degree of disc degeneration, in order to measure metabolic alterations and make similar metabolite change to be associated, can suitable amendment and repacking to be carried out to these HR-MAS tool and methods.

HR-MAS spectrum is used to measure the intervertebral disc across Tang Pusen rate range, to identify the compound that can carry out NMR observation, and determines the rate of change of these compounds between the different intervertebral disc degeneration stages.Relaxation time is also measured, to characterize the change of intervertebral disc chemical composition environment along with intervertebral disc degeneration and the degree of freedom of these chemical component.

2. materials and methods

A. the acquisition organized

This research have passed Ethics Committee's examination & approval.Lumbar vertebra obtains (the range of age: 22 ~ 85 years old) by surgical operation from n=17 tool people corpse, and-80 DEG C frozen.Vertebra autopsy saw and the dissecting knife of results dissociate.Reject the bone around intervertebral disc in vertebra.The biopsy punching of 3mm is carried out in the fibrous ring of intervertebral disc peeled off and nuclear area.These punchings to each other closely, and cylindrical in symmetrically.Average quality is 15.2 ± 3.4mg.Three of given area samples are side by side for measuring the reproducibility of wave spectrum.The general Lignum Rhamnellae level of soup is consistently read under the prerequisite strictly observing Tang Pusen staging.Have studied altogether general gloomy 1 grade of 8 soup, general gloomy 3 grades of 6 soup, general gloomy 5 grades of 6 soup.

The collection of B.HR-MAS data

HR-MAS data acquisition conditions: 1.0 ± 0.5 DEG C, rotating speed 2,250Hz, uses Varian (Varian) INOVA spectrometer, magnetic field intensity 11.75T ( ¹h, 500Hz), be equipped with 4mmgHX nanoneedle and visit.One dimension spectrum collection 40,000 complex point, spectrum width 20,000Hz (40ppm), pulse is wide 90 °, HOD presaturation phase 2s, 32 rank conversion, cycle period 8s (the longest T that > is 5 times ₁relaxation time), acquisition time 2s (the longest T that > is 5 times ₂relaxation time), total acquisition time 3:36min.Sample adopts 18 μ l zirconium rotors of traditional design to analyze, and comprises an oval sample room, the spiral top plug Leakage prevention of a sealing.After tissue sample is weighed and added rotor, each sample is drawn 3 μ 1 respectively and is added rotor bottom containing the heavy water (D2O+TSP, Sigma-Aldrich company) of the deuterated propanoic acid of 3-(trimethyl silyl)-2,2,3,3-tetra-of 0.75wt%.

Longitudinally (T ₁) relaxation time measurement adopt upset recover pulse train, time delay, excursion was 0.01 ~ 2.00s.Laterally (T ₂) measurement of relaxation time make use of rotor-synchronous (that is τ delay=n × (rotating speed) ^-1, n is even number) and Carr-Purcell-Meiboom pulse train, echo time excursion is 10 ~ 128ms.Compose for two-dimentional TOCSY, radial dimension (F2) spectrum width is 20,000Hz, obtains 4096 complex points; Indirect dimension spectrum width is 6,500Hz, obtains 256 complex points.TOCSY composes acquisition condition: 2s HOD presaturation/relaxation delay, acquisition time 0.2s, 32 steady-state pulses (only increasing by first), 16 instantaneous/cumulative, incorporation time excursion 10 ~ 80ms, phase sensitivity adopts States-Habercom, and total experimental period is about 5 hours 12 minutes.For making B ₀and B ₁heterogeneity effect minimizes, and adopts rotor-adiabatic instrument WURST-8 adiabatic pulses (33) of synchronous weighing apparatus to carry out isotropism mixing, Pulser apparatus (Pbox, Varian) adopts " Pandora ' sBox ", B ₁field is 6,5000Hz, and the persistent period is 444ms (1/ rotating speed).One dimension wave spectrum is obtained to assess the degraded of metabolite before and after each two dimension experiment.T ₁and T ₂the measuring object of relaxation time is vertebral pulp (n=9) and the fibrous ring (n=12) of intervertebral disc that is healthy and that degenerate.

C. date processing

Data are by VarianVNMR6.1C software (Varian Associates, Inc. (US) 611 Hansen Way, Palo Alto, California 94303, U.S.A., Palo Alto) online treatment, or utilize ACD/Labs 1D and 2D NMR process software 7.0 version (Advanced Chemistry Development company limited, Toronto) processed offline.The exponential function correction of one dimension FID signal, linearly widens the inverse that the factor equals acquisition time.Fourier transformation, phasing, makes TSP chemical shift be 0.00ppm.The calculating utilization index least-square analysis of relaxation time.This relaxation time is just used when only having the standard deviation when minimum power to be less than 10%.TOCSY data are at F ₁in utilize 3 × N linear prediction, be filled into 1024 complex points (only to F1) from 0, every one dimension all utilizes the correction of Gauss's weight.

Intersect the ownership at peak with reference to the chemical displacement value of bibliographical information before.On data visual evaluation basis, the following three Ge Pu districts of ownership of one-dimensional data: N-acetyl group (N-acetyl) district (1.90-2.10ppm); Choline head group (Cho) district (3.15-3.30ppm); And carbohydrate (Carb) district (3.50-4.20ppm).Three ratios, are abbreviated as N-Acetyl/Cho, Chol/Carb and N-Acetyl/Carb, after carbohydrate district integral area is decided to be 1.00, calculate three ratios of each wave spectrum.For the general Lignum Rhamnellae level of each soup, calculate average ratio and standard deviation, then carry out Student ' s t test to determine the statistical significance of data, only have and just think meaningful as p value < 0.05.

3. result

A. the difference of the general Lignum Rhamnellae level of soup

Soup general Lignum Rhamnellae level is respectively the annulus fibrosus disci intervertebralis district of 1,3,5 and the representative one dimension magic angle spinning (1-D HR-MAS) of nuclear area is composed as Figure 1-3.

Fig. 1 X, 1Y, 1Z show the representational 1-D HR-MAS obtained from the intervertebral disc (Fig. 1 X) of general gloomy 1 grade of soup, annular area (Fig. 1 Y) and nuclear area (Fig. 1 Z) and compose.Arrow shows the relation shown in Fig. 1 Y, 1Z between different collection of illustrative plates, represents position respective in intervertebral disc () shown in Fig. 1 X respectively.Circle represents the exemplary position of the 3mm biopsy perforation obtained from intervertebral disc.Distinguishable peak comprises: A: isoleucine, leucine, valine; B: lactate, isoleucine; C: alanine; F: the N-Acetyl resonance of chondroitin sulfate; G: glutamic acid; J: lysine; K: gallbladder alkali; L: phosphocholine; Square brackets district represents the C-H resonance of chondroitin sulfate.

Tang Pusen 1 grade of disc material with the ring of stiff easy bending and moisture gel core for feature (Fig. 1 X).The biopsy punching of 3mm obtains from the ring district of intervertebral disc and core district respectively, and Fig. 1 Y, 1Z respectively show its corresponding HR-MAS spectrum.Two HR-MAS spectrums all show to there is the resonance centre of N-acetyl at 2.04ppm, are connected to the resonance of the carbohydrate of chondroitin sulfate in 3.5 ~ 4.0ppm region.In addition, have also been observed Lactated resonance (1.33ppm), the resonance (ppm) of lipid, the resonance (3.21-3.25ppm) of choline head group, and several amino acid whose resonance (alanine (1.49ppm), isoleucine, leucine, and valine).What is interesting is, in the junior intervertebral disc of Tang Pusen, compared with ring district (Fig. 1 Y), core district (Fig. 1 Z) can observe lasting higher spectral resolution.

Fig. 2 X, 2Y, 2Z show the general gloomy 3 grades of intervertebral disc (Fig. 2 X) of the medium soup deteriorated, and corresponding HR-MAS spectrum derives from annular area (Fig. 2 Y) and nuclear area (Fig. 2 Z) respectively.Be the dehydration of intervertebral disc and the mechanical injuries of adjoint intervertebral disc substrate with the general gloomy 3 grades of relevant metamorphosis of soup.From spectroscopy, compared with the spectrum (Fig. 1 X, 1Y) of the general gloomy 1 grade of intervertebral disc of soup, the spectrally resolved property of the carbohydrate in the gloomy 3 Ji He districts of Tang Pu and ring district improves.The resonance (3.21ppm) comprising choline head group also enhances.

From pathology, Tang Pusen 5 grades of intervertebral disc (Fig. 3 X) show that intervertebral disc dewaters further, mucoprotein infiltration occurs and ring district decomposes on a large scale, and fibrous tissue replacement vertebral pulp, the visual difference between ring district and core district is reduced.From spectrally, the resonance in choline and carbohydrate district improves all further at wave spectrum Medium resolution and intensity.The reduction of N-Acetyl resonance intensity and the resonance number caused because of the generation of free amino acid and intensity can also be observed increase simultaneously.

Table 1 shows fibrous ring (A) and vertebral pulp (B) integration ratio and Student ' s t test result.Respective and average ± sdev (standard deviation) N-Acetyl/Cho, Cho/Carb and N-Acetyl/Carb ratio of Tang Pusen 1 grade, 3 grades, 5 grades intervertebral disc has all carried out statistics and has compared.Because Tang Pusen hierarchy rules is with subjectivity, so this research eliminates the intervertebral disc of the general Lignum Rhamnellae level 2 and 4 of soup.Concerning core district and ring district, N-Acetyl/Cho and Cho/Carb average ratio all show obvious difference in three soup general Lignum Rhamnellae levels, but N-Acetyl/Carb only exists significantly different general gloomy 3 grades of soup from the core district of 5 intervertebral disc.

Appoint ring district or core district and get two grades and compare, 1 grade of ratio 1 when comparing with 3 grades and ratio 2 are significant.Just think that comparing 1 grade is only significant with N-Acetyl/Carb during 5 Ji He district.N-Acetyl/Cho=integration (1.90-2.10ppm)/integration (3.15-3.30ppm), Cho/Carb=integration (3.15-3.30ppm)/integration (3.50-4.20ppm), N-Acetyl/Carb=integration (1.90-2.10ppm)/integration (3.50-4.20ppm).

In Fig. 4 A-4D, N-Acetyl/Cho (Fig. 4 A and 4B) and the Cho/Carb (Fig. 4 C and 4D) of three soup general Lignum Rhamnellae Ji Huan district (top) and core district (bottom) are divided out, to evaluate the overlapping cases between single ratio.Concerning N-Acetyl/Cho ratio, do not have overlapping between Tang Pu gloomy 1 grade and the single ratio in the core district of 5 intervertebral disc, and with have between other single ratio larger overlapping.To Cho/Carb ratio, between the single ratio in three soup general Lignum Rhamnellae Ji Huan districts, there is no overlap, do not have overlapping between Tang Pu gloomy 1 grade and the single ratio in the core district between 5.

B. total correlation spectrum (TOCSY)

In order to belong to the resonance observed in the one dimension spectrum of proton, two-dimentional TOCSY be obtained and compose, and the chemical shift at the intersection peak observed and the chemical displacement value previously reported are compared.Fig. 5 A shows the TOCSY spectrum of the intervertebral disc of gloomy 1 grade of Tang Pu.In the research that the general gloomy 1 grade of intervertebral disc of all eight soup is carried out, only observed the intersection peak of limited quantity, comprise the peak of the carbohydrate portions belonging to alanine (1.49,3.79ppm), lactate (1.35,4.16ppm) and proteoglycan aggregates.On the contrary, the TOCSY spectrum of six that the studied general gloomy 5 grades of intervertebral disc (Fig. 5 B) of soup degenerated then shows much more intersection peak that can detect, comprise isoleucine (0.92,1.32ppm), leucine (0.98,1.72ppm), lysine (1.73,3.04ppm), proline (1.73,1.93ppm), glutamine (2.14,2.46ppm and 2.14,3.79ppm), glutamic acid (2.1,2.36ppm), hydroxyproline (2.45,3.45ppm), and cholamine (3.15,3.83ppm).TOCSY experiment shows, the distinguishable resonance that occurs of carbohydrate district (3.5-4.2ppm) that 1-D HR-MAS composes be by several amino acids, containing cholamine complex and decompose the complex spike of the saccharide C-H proton got off from chondroitin sulfate.The 1-D HR-MAS of the intervertebral disc of all research compose to have at 3.21ppm and 3.23ppm place two unimodal, the chemical shift of the corresponding choline (Cho) that dissociates and phosphocholine (PC) respectively.TOCSY experiment also show Cho at the intersection peak of 3.55 × 4.07ppm and PC at the methene proton of 3.62 × 4.18ppm.Also have other little and wide resonance several also to identify in the choline district of wave spectrum.

C.T ₁and T ₂relaxation time

Fig. 6 A and 6B respectively illustrates the average T of the distinguishable resonance of gloomy 1 grade, 3 grades and the 5 grades intervertebral disc of Tang Pu ₁and T ₂relaxation time.In the collection of illustrative plates in all soup general Lignum Rhamnellae Ji He district and ring district, distinguishable resonance is only selected to carry out T ₁and T ₂the mensuration of relaxation time.So determine T to the resonance at 0.9,1.32,1.49,2.04,3.21 and 3.67ppm place ₁and T ₂relaxation time.The ring district determined and the T in core district ₁and T ₂relaxation time does not observe obvious change, therefore in conjunction with the relaxation time in these regions to increase the statistical significance compared between intervertebral disc grade.To the average T of the resonance of the general Lignum Rhamnellae level of each soup ₁and T ₂mensuration shows, along with intervertebral disc is constantly degenerated, and larger T ₁and T ₂change does not make metabolite produce obvious variation tendency.But along with the increase of the general Lignum Rhamnellae level of soup, can T be observed ₁relaxation time changes trend.T ₁the observable variation tendency of value is, T during most first arrival soup general gloomy 3 grades of intervertebral disc ₁value reduces, then T during gloomy 5 grades of intervertebral disc general to soup ₁value increases again.For T ₂, in observable resonance, there is no consistent variation tendency; But along with the increase of the general Lignum Rhamnellae level of soup, N-acetyl group (2.04ppm); There is the trend increased in the resonance (3.67ppm) of choline (3.21ppm) and carbohydrate C-H.

4. discuss

The ring district of intervertebral disc and the proton HR-MAS of core district sample compose closely similar; both wave spectrums all show and mainly come from the N-Acetyl resonance of chondroitin sulfate and the resonance of carbohydrate in a large number, and containing choline complex, lipid/lactate and several amino acid whose resonance.For the general gloomy 1 grade of intervertebral disc of soup, due to the character of the similar colloid in core district, make the resolution of its wave spectrum unanimously higher than ring district.

The important results that this research is observed is: along with the increase of the general Lignum Rhamnellae level of soup, can observe obvious change in the proton HR-MAS of disc annulus district and core district sample composes.Particularly at wave spectrum 3.5-4.0ppm resonance region, the corresponding loss with this region " multicomponent ", the observable resonance quantity depending on grade increases and formant can come to a point along with the increase of grade.In addition, in the spectral domain of compound comprising choline, the signal intensity of resonance can increase, and N-Acetyl resonance can weaken relatively.

Similarly, along with the increase of the general Lignum Rhamnellae level of soup, quantity and the intensity at the intersection peak in the TOCSY spectrum of intervertebral disc also can increase.Because aminoacid is as the existence of hydroxyproline, proline, glycine, lysine, leucine, isoleucine, alanine, valine, glutaminase and glutamic acid and cholamine, it is the composition of collagen mostly, thus degenerate core district and ring district TOCSY spectrum in there is a lot of intersection peak.Mainly amino acid whose resonance in the one dimension proton high resolution Magic angle spinning spectrum of the general gloomy 5 grades of nucleus pulposuses of soup and fibrous ring.Observe that amino acid whose phenomenon mainly causes due to the decomposition of collagen, this has observed and has been in utilizing the NMR of metalloprotein enzymic digestion cartilage to study before this.By inference, metalloproteases (MMPs) destroys the collagen supporting intervertebral disc, is therefore considered to play key player in the degeneration of intervertebral disc.Because collagenous network and collagen helix are decomposed into the aminoacid forming them, so these amino acid whose resonance all become more obvious in a peacekeeping two dimension magic angle spinning spectrum.

Research has also found the metabolism evidence that chondroitin sulfate is degraded along with the general Lignum Rhamnellae level increase of soup.The foundation of this conclusion is: compared with general gloomy 1 grade of soup; resonate (3.5-4.0ppm) intensity and resolution of carbohydrate C-H in the proton high resolution Magic angle spinning spectrum of 3 grades and 5 grades of annulus fibrosus disci intervertebralises and vertebral pulp both increases, and N-acetyl group (2.04ppm) resonance of chondroitin sulfate can weaken relatively.Pass through before ¹h and ¹³c magic angle spinning spectrum is studied bovine nasal cartilage that is natural and enzymic digestion; result shows; the composition of N-Acetyl resonance, by N-Acetyl resonance initial in undigested cartilage, becomes the resonance comprising N-acetyl group and ispol peak after digestion.Along with the degeneration of intervertebral disc, the content of chondroitin sulfate can reduce.TOCSY research shows, in the 3.5-4.0ppm region of degenerated spinal disc, comprises several amino acids, containing the mixture of the complexity of cholamine compound and the C-H resonance of carbohydrate, also causes distinguishable resonance owing to existing.Belonging to completely of resonance therefore also need to study the degraded of human disc in the intervertebral disc spectrum of degenerating, and be associated with the biochemical analysis (such as, dimethylated methylene base orchid (DMMB) is analyzed) of chondroitin sulfate and the biochemical analysis of collagen (collagenase).

In the HR-MAS spectrum observed, N-Acetyl resonance changes, on the basis that choline and carbohydrate/amino acid region resonance change; N-Acetyl/Cho, Cho/Carb and N-Acetyl/Carb ratio is studied, to determine which ratio best embodies the difference between the general Lignum Rhamnellae level of soup.The average ratio of N-Acetyl/Cho and Cho/Carb all shows obvious difference between three soup general Lignum Rhamnellae levels, and wherein the overlap of Cho/Carb ratio between the single numerical value of three soup general Lignum Rhamnellae levels is minimum.

Cho/Carb ratio does not have overlap between three soup general Lignum Rhamnellae level fibrous ring collection of illustrative plates, and the overlap between three soup general Lignum Rhamnellae level vertebral pulp collection of illustrative plates is minimum.This point is particularly useful, because concerning spectroscopy in body, can not require the spectrum obtaining independent core district or ring district so under many circumstances.Advantageously: they have very close signal and signal to noise ratio relatively.

It is useful that N-Acetyl/Chol ratio also can be proved to be for disc degeneration spectroscopic measurement in body, because N-Acetyl resonance is peak maximum in Tang Pu gloomy 1 grade and 3 grades of intervertebral disc, in the wave spectrum of the general gloomy 5 grades of intervertebral disc of soup, be reduced to the level also little or more suitable with it with carbohydrate district than choline.In fact, based on the reduction situation of N-acetyl base peak relative to choline and carbohydrate region, easily general for the soup obtained from He Huan district of core district gloomy 5 grades of collection of illustrative plates and soup general gloomy 1 grade and 3 grades of collection of illustrative plates naked eyes can be distinguished and come.

In research before, in order to characterize intervertebral disc and cartilage degradation, researcheres are studied the spin lattice relaxation time of aqueous solution and spin spin relaxation time.More hydrate tissues is found the T having longer water ₁and T ₂, and they can reduce along with the degeneration of intervertebral disc, and this may be based on the event of the loss (14) of moisture in tissue.Do not report T before ₁and T ₂change and disc degeneration product and the general Lignum Rhamnellae level of soup link together.

In this research, to the T of intervertebral disc catabolite ₁and T ₂the measurement result of meansigma methods shows very large transmutability.But at T ₁still certain trend can be observed in relaxation time.T ₁this trend of value is, demonstrates downward trend during most general gloomy 3 grades of first arrival soup, then general to soup gloomy 5 grades time demonstrate again the trend of rising.The general gloomy 3 grades of intervertebral disc catabolite T of the soup observed ₁the initial decline of value may be because disc degeneration result in the forfeiture of moisture; T during subsequently general to soup gloomy 5 grades ₁increase may be such as because the catabolite mobility that discharges from Dan Baiduotang proteoglycan PG and collagen stroma strengthen therefore.

The all decomposition prods T observed ₂change in, all there is no obvious trend.To the T of the water Sum decomposition product of a large amount of degenerated spinal disc ₁and T ₂measure, the information more useful to the relaxation time understanding measurement in this research can be obtained.

5. brief summary

In a word, the wave spectrum obtained from nucleus pulposus district and annular area sample by proton high resolution Magic angle spinning is closely similar.Importantly, the change of the general Lignum Rhamnellae level of the soup along with intervertebral disc, can observe obvious visual change in the proton high resolution Magic angle spinning wave spectrum of ring district and core district sample.From quantitative angle; the area ratio at two kinds of metabolite peaks in collection of illustrative plates; namely the ratio of the ratio of N-Acetyl resonance region and choline resonance zone, choline resonance zone and carbohydrate resonance zone can be used for the intervertebral disc distinguishing the general Lignum Rhamnellae level of different soup, and overlap between the single ratio of these different brackets is also very little.The T of chemical composition in disc spectra ₁and T ₂relaxation time change does not reflect the change of the water relaxation time of degenerated spinal disc as report before.Along with increasing the weight of of degenerating, the change of the relaxation time of intervertebral disc chemical composition reflects the change along with the cases of dehydration change of the increase intervertebral disc of the general Lignum Rhamnellae level of soup and the degree of decomposition of Dan Baiduotang proteoglycan PG and collagen stroma.In the body reflected in nuclear magnetic resoance spectrum, the chemical change of morphological characteristic to detection and disc degeneration is very useful.

In addition, their entirety, also with reference to knowing clearly following documents, is incorporated to herein to be incorporated by reference by this research:

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Embodiment 2

1. introduction

Traditional evaluation pain, the formation method of disc degeneration only pay close attention to morphological criteria.But, between the phenomenon that morphology is observed and patients symptomatic contact and little, this is known.A target of this in vitro study is exactly utilize quantitative magic angle spinning nuclear magnetic resoance spectrum as instrument, the biochemical marker of the intervertebral disc sample that accurate characterization is gathered in the crops from patient's surgical operation.The wave spectrum of the intervertebral disc obtained in the intervertebral disc marrow nuclear excising operation that patient implements because of backache by this research, and contrast with reference to the wave spectrum of intervertebral disc, to identify the biochemical signals of the disc degeneration of pain, wherein comprise with reference to intervertebral disc the intervertebral disc obtained from the operation of Patients Undergoing Scoliosis Correction.

2. materials and methods

Spectroscopy data acquisition condition: 11.7T (500MHz), 1 DEG C, rotating speed 2,250Hz, is equipped with the Varian INOVA spectrometer of 4mm gHX nano-probe.Utilize 35 μ l rotors of conventional design to respectively from discogenic pain patient operation obtain intervertebral disc (n=6) tissue, early stage and/or late period spinal fusion Patients Undergoing Scoliosis Correction perform the operation in acquisition intervertebral disc (n=4) tissue be studied.The quantitative proton of tissue sample (average=14.28 ± 2.91mg) is composed with D2O+0.75%TSP (Sigma-Aldrish company, St.Louis, MO) as interior mark.Each tissue sample adopts the synchronous Carr-Purcell-Meiboom-Gill of spin-echo rotor (CPMG) pulse train (nt=128, at=2.0s, TR=5s, echo time=80ms).To lactate resonance (1.31ppm; bimodal), N-acetyl group (PG) resonance (2.04ppm on Dan Baiduotang proteoglycan PG; unimodal), and collagen decomposition district (col) resonance (3.30-4.00ppm) carries out analyzing to contrast intervertebral disc sample.The detailed spectral portion that Fig. 7 A and 7B shows has carried out further note to these regions.

3. conclusion

Fig. 7 A and 7B respectively illustrates the 80ms of discogenic pain patient ¹h CPMG composes the 80ms of (a) and skoliosis patient ¹h CPMG composes (b).Wherein, Dan Baiduotang proteoglycan PG N-Acetyl resonance (PG), lactate and collagen decomposition district (col) are labeled out.

For deformation patients, these backaches show obviously lower PG/ lactate ratio and PG/col ratio (p < 0.05; See Fig. 7).In addition, the information list that the experiment wave spectrum that table 2 shows to Fig. 7 A and 7B shows is relevant.

4. discuss

This experimental result shows, biochemical marker may be used for characterizing the process relevant to discogenic pain.Research before once reported pH value to Dan Baiduotang proteoglycan PG synthesis and the impact of whole body health.Along with the increase of lactate concentration, due to H free in solution ⁺increase the effective pH value in intervertebral disc composition material is reduced, and then hinder the synthesis of Dan Baiduotang proteoglycan PG.

With regard to this experimental result, before the direct cause effect relation between lactate concentration and pain not understand by people, do not obtain detailed biochemistry yet and explain.But according to method described herein and brief summary, we disclose lactate concentration and are providing the advantageous application in the statistics relation between itself and pain.Do not consider the physiology association mechanism between these two parameters, this represent the instrument of a quite useful diagnosis painful area.The system and method that this is highly profitable described herein make the instrument of Noninvasive to be associated with pain measurement factor in there is obvious benefit, not consider to be hidden in below these results specifically biological " cause and effect " chemistry or biology relation herein.But someone thinks, Lactated increase can stimulate the nerve fiber in the granulation tissue relevant to intervertebral disc rehabilitation.Those skilled in the art are in disclosure summary and other useful information bases, clinical sample research that can match to a large amount of and catagen phase guides, and it confirms the purposes in these fields and other field further, namely can identify, utilize spectral signatures to assess biochemical degeneration and the relation with discogenic pain.

Be below relevant references, their entirety be incorporated to herein to be incorporated by reference:

Keshari?KR，Zektzer?AS，Swanson?MG，Majumdar?S，Lotz?JC，Kurhanewicz?J.Characterization?of?intervertebral?disc?degeneration?by?high-resolution?magic?angle?spinning(HR-MAS)spectroscopy.Magn?Reson?Med?2005；53(3)：519-527.

Maroudas?A.The?Biology?of?the?Intervertebral?Disc.In：Ghosh?P，editor.The?Biologyofthe?Intervertebral?Disc.Volume?2.Boca?Raton：CRC?Press；1988.p?Ch.9.

Urban?JP，Smith?S，Fairbank?JC.Nutrition?of?the?intervertebral?disc.Spine2004；29(23)：2700-2709.

It should be noted that, on above-mentioned disclosed basis, NMR wave spectrum can be used to identify and characterize intercentrum material, to determine the degree of degeneration of corresponding intervertebral disc, particularly material and direct, portentous, the reproducible relation of intervertebral disc soup general Lignum Rhamnellae inter-stage.Therefore, this represent the significantly useful and aspect of relative broad range of the present invention.The specific embodiment be described below is relevant with the application of magic angle spinning wave spectrum (HR-MAS), and this embodiment demonstrates special advantage to the disc material of the transplanting observed in diagnostic environment.The characteristic aspect that another embodiment described below is distinguishing biological tissue in body also show special advantage, represents other broad aspect of the present disclosure as follows.This can be completed by following embodiment: the Magic angle spinning system being particularly suitable for using biopsy sample utilizing other types; Or according to instruction below, appropriate reconstruction is carried out to the tool and method that will use, thus use other to can be used for the nuclear magnetic resonance, NMR spectra system of patient.

By being separated the high signal peak relevant to diagnosis pain, as exemplifying in embodiment 2 brief summary above, when these specific targets move in clinical practice in the equipment directly acting on human body from the MAS MRI equipment of 11.7 teslas, be considered to expand very well, a rear equipment is such as the MRI equipment of typical especially 3 or 1.5 teslas in clinical diagnosis.This is specially adapted to following situation: extra use customized local coil with along interested region, a certain specific region of such as lumbar vertebra and generate higher local field.

Further again, it should be noted that, tissue samples also can obtain in patient body by the mode of such as biopsy, then enters in the laboratory equlpment that such as high field MRI machine is such, the MAS NMR of such as 11.7 teslas, according to illustrating here, the various system and methods described are diagnosed patient.

In addition, various typical compound and/or wherein specific component factors be described to the target to the non-invasive diagnostic organizing medical condition.It should be noted that these compounds " factor " may comprise the compound or their molecular structure or its part or a kind of metabolite, catabolite or the by-product relevant to a certain degree to the compound that this is identified that are identified.In addition, the disclosure also relate to diagnostic tool and method and produces with the information showing to organize specific feature.This feature can comprise such as pain respectively and self degenerate with organizing.Or, also can comprise relevant to this fisrt feature or have another second feature causal.Such as, the inflammatory reaction of pain nerve, relevant somatomedin, a certain type, etc., cause effect relation can be had with in pain and tissue deterioration or both.These features can be the features that directly show of information that embodiment of the present invention produces, but this may be derived further by the feature shown and pain or the degenerates relevant useful diagnosis added and conclusion.We also predict, pain and to degenerate can be the result do not connected each other that draws of diagnostic tool described herein and method and target, also can together with this result connects each other with target.And the system and method for the novelty described herein can also identify the degree of these features.This can derive further result and conclusion, such as, in the derived weave spatial relationship of these features, such as, the position of certain intervertebral disc level or intervertebral disc part (or other organizational structure), wherein more pain or degeneration are more serious for this part, relevant to periarticular, level or region.This location can be produced by these useful informations oneself, also can be identified by further treatment and analysis on this basis.

Be enough to draw in the degree of these embodiment particular systems and device, no matter whether the disclosure, further predictive of other correlation technique, be described in detail the details beyond these method desired use.Those of ordinary skill in the art openly can understand these relations according to entirety of the present invention.Similarly, no matter whether following method, further clearly predictive of respective system and equipment, illustrate these systems and equipment outside method illustrates.Aforesaid content is relevant to the explanation told about below, also relevant to claim below.Such as but not limited to, it should be noted that some function aspects (or combination described between each key element) talked about in the system or equipment claim provided at this, further predictive of the method for additional this function of embodiment; Predictive of the independence enforcement aspect of following expection, although it does not need to be applied to as restriction this specific aspect and relevant pattern and embodiment, unless clearly carried out such description.

Although description above comprises many details, and to should not be construed as be limit the scope of the invention, and merely provide the example of some preferred embodiment for the present invention.Therefore; be to be understood that; scope of the present invention contains other embodiment that will be apparent to those skilled in the art completely; and therefore protection scope of the present invention not limited by any scheme outside attached claim; when wherein mentioning separately certain key element; unless explicitly stated otherwise, otherwise not refer to " one and only have one " and refer to " one or more ".In above-mentioned preferred implementation known by above-mentioned those of ordinary skill, all structural, chemical, the function equivalent of each key element can specifically merge, and ought to be included in claim of the present invention.In addition, there is no need to state each problem that device of the present invention or method will solve, because it will comprise in detail in the claims.Whether in addition, no matter propose clearly in the claims, any key element, composition or method step disclosed by the invention be not all for declaration of will open to the public.Any key element of this claim do not explained by the 112nd article the 6th section, United States code the 35th chapter, unless this key element clearly uses word " to mean " statement.

Table 1A: intervertebral disc metabolite ratio, fibrous ring

Integral area

1 grade

3 grades

5 grades

1?vs?3

?1?vs?5

?3?vs?5

?N-Acetyl/Cho

?3.685±0.601

?2.552±0.339

?1.941±0.54.

0.006

?0.035

?0.035

?Cho/Carb

?0.115±0.027

?0.191±0.014

?0.243±0.016

＜0.001

?＜0.001

?＜0.001

?N-Acetyl/Carb

?0.420±0.137

?0.487±0.062

?0.466±0.096

0.277

?0.486

?0.638

Table 1B: intervertebral disc metabolite ratio, vertebral pulp

Integral area

1 grade

3 grades

5 grades

?1?vs?3

1?vs?5

?3?vs?5

?N-Acetyl/Cho

?5.487±1.400

?4.336±0.0.076

?2.364±0.411

?0.031

＜0.001

?0.001

?Cho/Carb

?0.082±0.018

?0.121±0.014

?0.161±0.006

?0.00l

＜0.001

?0.001

?N-Acetyl/Carb

?0.448±0.139

?0.515±0.049

?0.380±0.079

?0.465

0.146

?0.008

Table 2: ratio PG/ lactate, PG/Col, lactate/Col+ standard deviation

?	Discogenic pain	Skoliosis
			PG/ lactate PG/Col lactate/Col	0.37±0.36 0.28±0.14 0.70±0.35	1.72±0.81 0.66±0.35 0.3?8±0.08

Claims (14)

1. a medical diagnostic system, it provides the useful information of the degree of the feature of the first area of instruction disc tissue, and wherein this feature is relevant to pain, and this system comprises:

Nuclear magnetic resonance, NMR (NMR) spectra system, it can provide the NMR of the chemical composition of the first area inner tissue of tissue to compose, and provide and compose relevant data to this NMR, this data format is suitable for the useful information of the feature being processed into the first area that can indicate tissue; And

A kind of processor, for the treatment of the data obtained in the NMR spectrum of the first area from tissue, and exports the numerical value of the parameter relevant with the analysis that pain is correlated with, thus provides useful information by the data processed.

2. system according to claim 1, is characterized in that, described system is for the degree of the feature of the second area of the degree and contiguous first area that compare the feature of first area.

3. system according to claim 1, is characterized in that, described first area comprises nucleus pulposus part.

4. system according to claim 3; it is characterized in that; described feature is also relevant to tissue deterioration, and the evaluation of described tissue deterioration is at least partly based on calculating at least one organizing in the following chemical components ratio of first area described: N-acetyl group compares carbohydrate than choline and choline.

5. system according to claim 3, is characterized in that, described feature is also relevant to tissue deterioration, and the evaluation of described tissue deterioration is at least partly based on the described chondroitin sulfate organizing first area.

6. system according to claim 3, it is characterized in that, described feature is also relevant to tissue deterioration, and the degree of described tissue deterioration at least comprises one of following: the degree of decomposition of described intervertebral disc dehydration degree, the proteoglycan matrix relevant to described intervertebral disc and the degree of decomposition of collagen stroma of being correlated with described intervertebral disc.

7. system according to claim 3, it is characterized in that, described first area comprises two chemokineses relevant to pain, and described useful information is at least partly based on the numeric ratio of at least one measurement parameter, wherein this parameter is relevant to each in described two chemokineses in first area.

8. system according to claim 7, is characterized in that, one of described two chemokineses are proteoglycan-related factor.

9. system according to claim 7, is characterized in that, one of described two chemokineses are lactate-related factor.

10. system according to claim 8, is characterized in that:

Described proteoglycan-related factor comprises N-acetyl group.

11. systems according to claim 7, is characterized in that, one of described two chemokineses are collagen-related factor.

12. systems according to claim 11, is characterized in that, described collagen-related factor comprises the chemical entities of instruction collagen decomposition.

13. systems according to claim 1, is characterized in that,

The described NMR spectrum obtained from the first area of described tissue comprises lactate-related resonance and proteoglycan-related resonance, described processor also for analyzing lactate-related resonance and proteoglycan-related resonance according to the described NMR spectrum obtained from the first area of described tissue, thus produces the numerical value of described relevant with the analysis that pain is correlated with parameter.

14. systems according to claim 8, is characterized in that, the another one in described two chemokineses is lactate-related factor.