CN105708861B - The new opplication of the excretion body of source for mesenchymal stem cells - Google Patents
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Abstract
The invention discloses application of the excretion body of source for mesenchymal stem cells in the drug of preparation treatment ankylosing spondylitis.The excretion body of source for mesenchymal stem cells has inhibiting effect to chronic inflammation and pathologic skeletonization, and has the advantages that side effect is low, at low cost, effect is more longlasting.
Description
Technical field
The present invention relates to the outer of the purposes of the excretion body of source for mesenchymal stem cells more particularly to source for mesenchymal stem cells
Secrete application of the body in the drug of preparation treatment ankylosing spondylitis.
Background technique
Ankylosing spondylitis (abbreviation AS) is a kind of common autoimmune disease, and illness rate is higher, China about
For 0.2~0.54% (reference can be made to: Rong J, Jieruo G.Spondyloarthritis in China.Current
opinion in rheumatology.2013；25(4):460-7.).AS is apt to occur in young man, often involves axis Bones and joints,
Major Clinical and pathological characters be chronic progressive inflammation and pathologic skeletonization (reference can be made to: Braun J, Sieper
J.Ankylosing spondylitis.Lancet.2007；369(9570):1379-90.).However, the pathogenesis mesh of AS
Before be still not clear, relationship of the two big pathological characters-between inflammation and skeletonization is also indefinite, and pathogenesis is unclear to be caused at present
AS treatment means limitation is big, validity is low.With the progress of disease, AS patient will gradually lose life and labour capacity, give
Patient brings heavy psychological pressure and financial burden, causes tremendous influence and obstruction to the development of society.Therefore, it visits
Suo Jineng alleviates AS patient's body inflammation can prevent the treatment method of pathologic skeletonization particularly important again.
The treatment means of AS include non-drug therapy, drug therapy and three kinds of operative treatment [15,16] at present.Non-drug is controlled
Treat mainly includes patient education, reconditioning, physiotherapy etc..Drug therapy includes non-steroid anti-inflammatory drug (such as Diclofenac
Sodium, celecoxib, loxoprofen etc.), alleviate state of an illness antirheumatic drug (such as sulfasalazine, methotrexate (MTX)), glucocorticoid
(such as corticosteroid), antalgesic (such as brufen) and biological agent (such as tumor necrosis factor antagonists) etc..Operation
Treatment includes replacement of total hip, spinal osteotomy orthopaedy etc..
In terms of the treatment of AS, non-drug therapy is mainly used for assisting in the treatment of, and the final state of an illness that can not change AS patient turns
Return.Drug treatment, though the pain symptom and level of inflammation of non-steroid anti-inflammatory drug, antalgesic energy reduction of patient, nothing
Method improves the pathologic bon e formation of patient, also have the side effects such as gastrointestinal reaction, cardiovascular response (reference can be made to: Miceli-
Richard C,Dougados M.NSAIDs in ankylosing spondylitis.Clinical and
experimental rheumatology.2002；20(6Suppl 28):S65-6.)；Alleviate state of an illness antirheumatic drug to be only capable for the treatment of
The periphery arthritic symptom of AS patient, be unable to improve axis inflammation of bone and pathologic skeletonization (reference can be made to: van der Heijde
D,Sieper J,Maksymowych WP,Dougados M,Burgos-Vargas R,Landewe R,et al.2010
Update of the international ASAS recommendations for the use of anti-TNF
agents in patients with axial spondyloarthritis.Annals of the rheumatic
diseases.2011；70(6):905-8.)；Corticosteroid is simply possible to use in locally injecting and relieves pain symptom, for whole body inflammation
Disease level and pathologic ossification also do not act on；Biological agent treatment, especially tumor necrosis factor antagonists are treated, and are current
One of main treatment method of AS, though studies have found that it can inhibit AS patient's body inflammation, but can not prevent even
Backbone pathologic bon e formation can also be accelerated, furthermore biological agent high medical expense, some patients are difficult to undertake, and also have sense
The side effects such as dye, tumour, hepatic and renal function exception (reference can be made to: van der Heijde D, Landewe R, Einstein S, Ory
P,Vosse D,Ni L,et al.Radiographic progression of ankylosing spondylitis after
up to two years of treatment with etanercept.Arthritis and rheumatism.2008；58
(5):1324-31.).In terms of operative treatment, it is mainly used for the treatment such as advanced stage AS patient spine deformity, hip lesion, it can not
Reach the target for curing AS.
Mescenchymal stem cell (mesenchymal stem cells, MSCs) is derived from the non-hematopoiesis system of mesoblastic one kind
System multipotential stem cell has confirmed that MSCs in addition to self-renewing and multi-lineage potential, also has very strong anti-inflammatory and suppression
The ability of panimmunity cell processed, and peripheral immune tolerance can be induced.Research shows that MSCs is adjusted by secretion panimmunity
The factor, such as IFN-γ, PGE2 (reference can be made to: Polchert D, Sobinsky J, Douglas G, Kidd M, Moadsiri
A,Reina E,et al.IFN-gamma activation of mesenchymal stem cells for treatment
and prevention of graft versus host disease.European journal of
immunology.2008；38 (6): 1745-55. and Spaggiari GM, Abdelrazik H, Becchetti F,
Moretta L.MSCs inhibit monocyte-derived DC maturation and function by
selectively interfering with the generation of immature DCs:central role of
MSC-derived prostaglandin E2.Blood.2009；113 (26): 6576-83.), to inhibit immunocyte (such as
T cell, B cell, NK cell, antigen presenting cell etc.) function (reference can be made to: Corcione A, Benvenuto F,
Ferretti E,Giunti D,Cappiello V,Cazzanti F,et al.Human mesenchymal stem cells
modulate B-cell functions.Blood.2006；107 (1): 367-72. and Di Nicola M, Carlo-
Stella C,Magni M,Milanesi M,Longoni PD,Matteucci P,et al.Human bone marrow
stromal cells suppress T-lymphocyte proliferation induced by cellular or
nonspecific mitogenic stimuli.Blood.2002；99(10):3838-43.).
Excretion body is a kind of diameter secreted by cell about 30~100nm, and density range is between 1.13~1.19g/ml
Film property microcapsule bubble.Excretion body can carry a variety of protein similar with source cell, mRNA, miRNA, participate in immunological regulation, thin
The processes such as born of the same parents' communication, cell migration, angiogenesis (reference can be made to: Yanez-Mo M, Siljander PR, Andreu Z, Zavec
AB,Borras FE,Buzas EI,et al.Biological properties of extracellular vesicles
and their physiological functions.Journal of extracellular vesicles.2015；4:
27066.).The excretion body in the discovery such as Lai source MSCs can reduce myocardial ischemia-reperfusion injury, and confirm excretion body
MiRNA can promote angiogenesis, thus excretion body be likely to become treatment cardiovascular disease a new direction (reference can be made to: Lai
RC,Arslan F,Lee MM,Sze NS,Choo A,Chen TS,et al.Exosome secreted by MSC
reduces myocardial ischemia/reperfusion injury.Stem cell research.2010；4(3):
214-22.).Xin etc. has found that the excretion body in the source MSCs by shifting miR-133b to nerve cell, can promote neural axon
Growth (reference can be made to: Xin H, Li Y, Buller B, Katakowski M, Zhang Y, Wang X, et al.Exosome-
mediated transfer of miR-133b from multipotent mesenchymal stromal cells to
neural cells contributes to neurite outgrowth.Stem cells.2012；30(7):1556-
64.).The excretion body in the discovery such as Filipazzi tumour cell source can pass through NK cell-stimulating receptor NKG2D (natural-
Killer group 2, member D) inhibit T cell and NK cell cytotoxicity, so that the immune system for influencing host (can
Referring to: Filipazzi P, Burdek M, Villa A, Rivoltini L, Huber V.Recent advances on the
role of tumor exosomes in immunosuppression and disease progression.Seminars
in cancer biology.2012；22(4):342-9.).
In terms of using the excretion body in the source MSCs treatment disease, there are two patents, Kiang LS etc. uses the source MSCs
Excretion body treatment mouse pulmonary fibrosis, can significantly improve mouse survival rate, mitigate bleomycin (bleomycin, BLM) and lead
Cause pulmonary fibrosis pulmonary lesion (reference can be made to: Kiang LS, Wei YMS, Sheng CT, Chai LR.USE OF EXOSOMES
TO PROMOTE OR ENHANCE HAIR GROWTH patent US2015024011.2015-01-22.).Liu Ming etc. will
The excretion body in the source MSCs for promoting wound repair and hair growth, obtain good result (reference can be made to: Liu Ming, Xu Jun,
Inventors mescenchymal stem cell excretion body applies patent in the pharmaceutical preparation of preparation treatment pulmonary fibrosis
CN104666344A.2015-06-03.)。
But up to the present, at home and abroad there is no the reports and patent of the excretion body treatment AS using the source MSCs.
Summary of the invention
Source for mesenchymal stem cells is provided it is an object of the invention to overcome the shortcomings of the prior art place
Application of the excretion body in the drug of preparation treatment ankylosing spondylitis.The excretion body in the source MSCs is to chronic inflammation and pathologic
Skeletonization has inhibiting effect, and has the advantages that side effect is low, at low cost, effect is more longlasting.
As further improvement to above-mentioned technical proposal, the mescenchymal stem cell is mesenchymal stem cell.
As further improvement to above-mentioned technical proposal, the mesenchymal stem cell is prepared by following methods
Arrive: then the separation and Extraction mescenchymal stem cell from marrow is obtained through originally culture and secondary culture.
As the further improvement to above-mentioned technical proposal, the extracting method of the excretion body are as follows: cultivate enough bones
Bone marrow-drived mesenchymal stem, the 48h before extracting stem cell excretion body clean cell with PBS, replace serum free medium, continue to train
Support 48h；
Serum free medium is collected, 300 × g is centrifuged 10min, takes supernatant；2000 × g is centrifuged 10min, takes supernatant；10000
× g is centrifuged 30min, takes supernatant；100000 × g is centrifuged 70min, takes precipitating；Precipitating is resuspended in PBS, and 100000 × g is centrifuged 70min,
Collection precipitates up to the excretion body.
As further improvement to above-mentioned technical proposal, the concentration of the excretion body in the drug is 1.3 × 109~
3.5×109Grain/milliliter.
As further improvement to above-mentioned technical proposal, the drug also includes pharmaceutically acceptable carrier." pharmacy
Upper acceptable carrier " includes the volumes of any and all physical compatibilities, decentralized medium, coating material, antibacterium and antimycotic
Agent, etc. blend and absorb delayer etc..The example of pharmaceutically acceptable carrier includes water, salt water, phosphate buffered saline, dextrose
One or more of acid anhydride, glycerol, ethyl alcohol etc. and their combination.In many cases it is preferred to be by isotonic agent for example
Sugar, polyalcohol or sodium chloride include in the composition.Pharmaceutically acceptable carrier also may include it is a small amount of can improve antibody or
The shelf life of antibody moiety or the auxiliary substance of validity, such as wetting agent or emulsifier, preservative or buffer.
Drug of the invention can be diversified forms.These forms include such as liquid, semisolid and solid dosage forms, such as liquid
Liquid solution agent (such as injectable and infusible solutions agent), dispersing agent or suspension, tablet, pill, powder, liposome and bolt
Agent.Preferred form depends on scheduled administration mode and treatment use.Preferably, the pharmaceutical dosage form is injection or freeze-drying
Preparation.Typically preferred drug is injectable or infusible solutions dosage form formula.Preferred administration mode is parenteral (such as vein
In interior, subcutaneous, peritonaeum, intramuscular) administration.In a preferred embodiment, drug of the invention passes through intravenous infusion
Or injection is given.In another preferred embodiment, drug of the invention is given by intramuscular or subcutaneous injection.May be used also
Supplementary active compounds are incorporated into drug, in certain embodiments, can by drug of the present invention and it is one or more can
Other healing potion for treating AS is prepared altogether together and/or is given altogether.For example, can be by drug of the invention and a kind of or more
Kind can be prepared altogether together and/or be given altogether in conjunction with the drug of other targets.
We have found that the excretion body in the source MSCs can obviously inhibit the proliferation of T cell and CTL thin in the research of early period
Cytotoxicity, induction Th1/Th2 is balanced to be tilted to Th2, and cause Treg generates.By being injected intravenously to AS patient
The excretion body in the source MSCs, it has been found that it has the function of being relieved AS patient's inflammation and backbone ossification.It proves for the first time, benefit
With the excretion body in the source MSCs, treatment AS patient's body chronic inflammation and the big pathologic problems of pathologic skeletonization two can be reached simultaneously
Effect.This is a kind of completely new, effective AS treatment method.
Compared with existing treatment technology, the present invention has the advantage that
1) the excretion body in the source MSCs has inhibiting effect to chronic inflammation and pathologic skeletonization.It proves, utilizes for the first time
The excretion body in the source MSCs can reach treatment AS patient's body chronic inflammation and the big pathologic problems of pathologic skeletonization two simultaneously
Effect.It can only delay relative to non-steroid anti-inflammatory drug, antalgesic, alleviation state of an illness antirheumatic drug, tumor necrosis factor antagonists etc.
The drug of inflammation is solved, the excretion body in the source MSCs has more fully function and effect.
2) low side effect.The AS therapeutic agent of the past is only capable of delaying disease progression to a certain extent, can not finally prevent disease
Journey, while there is also many adverse reactions and side effects.The excretion body immunogenicity in the source MSCs is low, treats AS patient, does not send out
Now infection, gastrointestinal reaction, cardiovascular response, tumour occur, a problem that hepatic and renal function is abnormal, prompt its side effect compared with
It is small.
3) inexpensive.The excretion body in the source MSCs can constantly be secreted by MSCs, relative to tumor necrosis factor antagonists etc.
Biological agent, cost is relatively low.
4) effect is more longlasting.The excretion body in the source MSCs has biological characteristics similar with MSCs, simultaneously because it is straight
Diameter only has about 40~100nm, is not easy to be retained by organs such as liver, lung, spleens, is easier to recycle in vivo, can play more longlasting
Therapeutic effect.
Detailed description of the invention
Fig. 1 is excretion body surface face Protein Detection figure；
Fig. 2 is treatment flow chart；
Fig. 3 shows that ankylosing spondylitis treats the facial symptom after preceding and treatment using excretion body with psoriatic,
The face for treating up till now patient covers with fash, and with pain, limitation of activity, these symptoms have obtained significant alleviation after treatment；
Fig. 4 shows that ankylosing spondylitis is improved with the articulatio sacroiliaca lesion of psoriatic；
Fig. 5 shows that ankylosing spondylitis is significantly dropped with the laboratory checking index of psoriatic with disease activity index
It is low.
Specific embodiment
Existing therapeutic scheme (including whole drug therapies as described above) is controlled for the chronic inflammation of AS
It treats, and attempts by inhibiting inflammation to delay pathologic skeletonization.Though these methods can alleviate inflammation, nothing to a certain extent
Method inhibit pathologic skeletonization, while there is also somewhat expensive, patient can not be resistant to, side effect is more the disadvantages of.
The present invention is different from existing treatment means, has not yet to see the case of the excretion body treatment AS with the source MSCs,
The treatment method can not only significantly inhibit inflammation, can also inhibit pathologic skeletonization, while without obvious toxic-side effects.
To better illustrate the object, technical solutions and advantages of the present invention, below in conjunction with specific embodiment to the present invention
It is described further.
Embodiment
The excretion body in the source MSCs is for treating AS chronic inflammation and pathologic skeletonization
(1) volunteer screens
20~30 one full year of life volunteers are chosen, project as required checks UP, and after qualified, 30ml marrow is extracted in reservation.
(2) separation, identification of M SCs
Isolated MSCs, progress cellular morphology and phenotypic evaluation from obtained marrow, and do microbiologic inhibition tests.
(3) MSCs is expanded, and excretion body, which extracts, to be prepared
By qualified MSCs amplification cultivation, according to 4 × 107The enough MSCs of cell concentration culture, it is outer extracting
48h before body is secreted, the cell culture fluid containing serum is removed, PBS is washed three times, and serum free medium is added, cultivates 48h.
(4) excretion body is extracted and is detected
Serum free medium is collected, 300 × g is centrifuged 10min, takes supernatant；2000 × g is centrifuged 10min, takes supernatant；10000
× g is centrifuged 30min, takes supernatant；100000 × g is centrifuged 70min, takes precipitating；Precipitating is resuspended in PBS, and 100000 × g is centrifuged 70min,
Collecting precipitating is excretion body.Part excretion body sample is taken, Electronic Speculum, surface markers, microbiologic inhibition tests are carried out.Excretion body surface face egg
White detection figure is as shown in Figure 1.
(5) AS patient enters group
It is all enter group case diagnosis meet ASAS formulation mesinae rigid spine inflammation diagnostic criteria (MRI is positive+extremely
Few 1 SpA sign or the HLA-B27 positive+at least two SpA signs), BASDAI >=4, spinal pain
VAS scoring >=4, between age requirement 18~40 years old, course of disease time is required from being made a definite diagnosis for the first time no more than 5 years, and chronic low back is bitterly
Symptom at least three moon, do not received biological agent treatment in all patient's courses of disease, and tumour markers check it is negative.
All patients in group sign informed consent form, receive the source MSCs excretion body infusion of therapeutic.
(6) excretion vena systemica is injected
AS patient is divided into three groups, including the source MSCs excretion body treatment group, tumor necrosis factor antagonists (benefit match is general) group
And control group.The source MSCs excretion body treatment group gave the treatment of venoclysis excretion body in the 1st~8 week, according to 1.3~3.5 ×
1010Excretion body particle total amount be unit (reference can be made to: Kordelas L, Rebmann V, Ludwig AK, Radtke S,
Ruesing J,Doeppner TR,et al.MSC-derived exosomes:a novel tool to treat
therapy-refractory graft-versus-host disease.Leukemia.2014；28 (4): 970-3.), it carries out
Intravenous injection, 2 times a week, totally 16 times；Tumor necrosis factor antagonists group gave general 25mg/ times subcutaneous note of benefit match in the 1st~8 week
Treatment is penetrated, 2 times a week, totally 16 times；Control group gave placebo venoclysis treatment, 2 times a week, totally 16 times in the 1st~8 week.
(7) therapeutic effect is assessed
Respectively at 0,1,2,3,4,5,6,7,8,16,24,32,40,48 week collection each group AS patients serum, detect ESR,
The indexs such as CRP, TNF-α, IL-6, IL-17, IL-23, to assess its current internal level of inflammation.MRI is checked, observes local ridge
Column, articulatio sacroiliaca pathologic skeletonization situation.
(8) interpretation of result
As the data in 1~table of table 8 are shown: compared to the control group, the source MSCs excretion body treatment group and tumor necrosis factor
Antagonist group can inhibit AS patient's body level of inflammation (ESR, CRP, TNF-α, IL-6, IL-17, IL-23 are significantly reduced),
And the source MSCs excretion body treatment group inhibits inflammation ability more stronger than tumor necrosis factor antagonists group.
Compared to the control group, the source MSCs excretion body treatment group significantly inhibits AS patient spine pathologic bon e formation, and tumour
Necrosis factor antagonists group does not influence (the pathologic ossification scoring: 0 point: normal of pathologic bon e formation progress；1 point: vertebral rim has
Slight hyperplasia；2 points: the obvious spur of vertebral rim is formed；3 points: side vertebral rim forms bone bridge；4 points: bilateral vertebral rim shape
Skeletonization bridge, intervertebral fusion).Treatment flow chart is as shown in Figure 2.Fig. 3~Fig. 5 is a wherein ankylosing spondylitis with psoriasis
The treatment condition figure of patient: Fig. 3 shows that this ankylosing spondylitis treats preceding and treatment using excretion body with psoriatic
Facial symptom afterwards, the face for treating up till now patient covers with fash, and with pain, limitation of activity, these symptoms are obtained after treatment
Significant alleviation is arrived；Fig. 4 shows that this ankylosing spondylitis is improved with the articulatio sacroiliaca lesion of psoriatic；Fig. 5
Show that this ankylosing spondylitis is significantly reduced with the laboratory checking index and disease activity index of psoriatic.Table 1
~table 8 is as follows.
Table 1: treatment number of cases and effective percentage
| Number of cases | Efficient (%) | |
| It is total | 89 | 68.5 |
| Male AS | 57 | 70.1 |
| Women AS | 32 | 65.6 |
| Early stage AS | 35 | 82.8 |
| Middle and advanced stage AS | 54 | 59.2 |
Note: during treatment effectively refers to the excretion body treatment in the source MSCs, Bath ankylosing spondylitis disease activity index is commented
Estimating (BASDAI) or ankylosing spondylitis state of an illness mobility assessment (ASDAS) reduces by 2 or more.
Table 2: erythrocyte sedimentation rate (ESR) (ESR) level (mm/h) compares
Table 3:C reactive protein (CRP) level (ng/ml) compares
| The source MSCs excretion body treatment group | Tumor necrosis factor antagonists group | Control group | |
| 0 week | 63.4±2.1 | 60.9±3.8 | 62.0±4.1 |
| 1 week | 61.2±3.9 | 59.4±5.2 | 64.1±1.3 |
| 2 weeks | 57.8±1.7 | 56.6±4.8 | 56.4±4.8 |
| 3 weeks | 45.2±4.7 | 56.1±4.0 | 62.2±2.7 |
| 4 weeks | 43.3±2.9 | 44.3±2.8 | 59.7±2.1 |
| 5 weeks | 32.5±1.8 | 37.5±6.7 | 59.2±7.9 |
| 6 weeks | 20.9±4.9 | 33.8±1.1 | 62.0±3.4 |
| 7 weeks | 17.6±4.5 | 28.4±3.3 | 57.4±0.6 |
| 8 weeks | 7.6±4.5 | 16.4±3.8 | 56.3±4.6 |
| 16 weeks | 9.2±2.2 | 20.2±1.9 | 58.6±2.8 |
| 24 weeks | 17.7±1.6 | 28.4±2.6 | 45.5±5.5 |
| 32 weeks | 27.7±5.3 | 45.3±5.2 | 62.3±1.5 |
| 40 weeks | 39.0±4.7 | 55.4±1.4 | 57.8±4.1 |
| 48 weeks | 44.4±2.5 | 65.7±2.9 | 67.5±2.6 |
Table 4:TNF- alpha levels (pg/ml) compare
| The source MSCs excretion body treatment group | Tumor necrosis factor antagonists group | Control group | |
| 0 week | 475.3±15.6 | 477.5±23.4 | 439.3±22.5 |
| 1 week | 453.1±25.2 | 481.6±15.5 | 401.6±32.0 |
| 2 weeks | 377.8±18.7 | 356.6±24.8 | 456.4±48.8 |
| 3 weeks | 245.2±45.7 | 296.1±47.0 | 382.2±24.7 |
| 4 weeks | 213.3±26.9 | 284.3±28.8 | 349.7±23.1 |
| 5 weeks | 196.8±11.9 | 247.5±12.2 | 380.3±22.3 |
| 6 weeks | 133.6±6.3 | 210.9±14.8 | 402.0±14.4 |
| 7 weeks | 109.3±13.8 | 185.3±13.9 | 369.2±15.8 |
| 8 weeks | 89.3±11.4 | 178.5±22.5 | 392.3±20.1 |
| 16 weeks | 258.2±8.9 | 333.7±32.0 | 382.1±35.9 |
| 24 weeks | 299.4±18.3 | 404.8±21.6 | 389.3±19.4 |
| 32 weeks | 259.7±43.6 | 463.8±34.1 | 467.3±52.0 |
| 40 weeks | 312.5±17.5 | 427.9±22.1 | 484.3±26.7 |
| 48 weeks | 418.4±25.9 | 524.7±28.0 | 419.8±29.6 |
Table 5:IL-6 horizontal (pg/ml) compares
Table 6:IL-17 horizontal (pg/ml) compares
| The source MSCs excretion body treatment group | Tumor necrosis factor antagonists group | Control group | |
| 0 week | 416.4±13.7 | 432.5±19.5 | 418.3±12.7 |
| 1 week | 374.2±24.9 | 399.7±37.8 | 455.4±35.4 |
| 2 weeks | 313.4±22.4 | 384.5±18.5 | 364.7±13.1 |
| 3 weeks | 293.5±13.9 | 346.5±22.3 | 484.2±12.4 |
| 4 weeks | 237.6±6.7 | 311.7±15.9 | 402.0±14.4 |
| 5 weeks | 207.2±16.5 | 264.7±29.0 | 395.3±32.7 |
| 6 weeks | 183.9±29.6 | 199.2±36.3 | 378.4±9.7 |
| 7 weeks | 75.7±26.2 | 148.6±32.8 | 373.7±13.6 |
| 8 weeks | 54.3±11.4 | 128.1±8.1 | 452.5±24.6 |
| 16 weeks | 194.6±14.8 | 223.5±12.1 | 389.2±34.4 |
| 24 weeks | 239.7±23.6 | 289.7±19.5 | 345.6±29.3 |
| 32 weeks | 229.3±9.8 | 333.6±39.7 | 395.5±22.1 |
| 40 weeks | 334.9±26.8 | 368.1±12.4 | 372.6±17.4 |
| 48 weeks | 388.1±14.5 | 445.1±14.3 | 415.7±24.6 |
Table 7:IL-23 horizontal (pg/ml) compares
Table 8: the scoring of backbone pathologic bon e formation is compared
| The source MSCs excretion body treatment group | Tumor necrosis factor antagonists group | Control group | |
| 0 week | 4.2±1.9 | 4.7±2.8 | 4.4±1.4 |
| 1 week | 4.6±1.5 | 4.2±1.9 | 4.3±0.6 |
| 2 weeks | 4.7±2.0 | 4.7±2.3 | 5.3±1.5 |
| 3 weeks | 5.0±0.4 | 5.1±2.9 | 4.6±1.4 |
| 4 weeks | 5.4±1.5 | 5.2±0.7 | 4.6±0.2 |
| 5 weeks | 4.3±1.1 | 4.2±1.9 | 5.5±1.7 |
| 6 weeks | 4.0±0.7 | 4.6±1.2 | 4.5±1.7 |
| 7 weeks | 4.1±1.6 | 5.3±1.5 | 5.4±0.6 |
| 8 weeks | 3.8±1.7 | 4.4±1.6 | 4.5±1.6 |
| 16 weeks | 4.0±0.2 | 4.5±0.7 | 4.4±1.5 |
| 24 weeks | 3.5±0.8 | 3.9±1.3 | 4.9±1.4 |
| 32 weeks | 2.8±1.3 | 4.8±1.1 | 4.7±1.7 |
| 40 weeks | 2.2±0.3 | 4.8±2.1 | 5.8±2.8 |
| 48 weeks | 2.9±0.2 | 5.3±3.6 | 5.3±1.6 |
Finally, it should be noted that the above embodiments are merely illustrative of the technical solutions of the present invention rather than protects to the present invention
The limitation of range is protected, although the invention is described in detail with reference to the preferred embodiments, those skilled in the art should
Understand, it can be with modification or equivalent replacement of the technical solution of the present invention are made, without departing from the essence of technical solution of the present invention
And range.
Claims (7)
1. the excretion body in mesenchymal stem cell source answering in the drug of preparation treatment ankylosing spondylitis companion's psoriasis
With.
2. application according to claim 1, it is characterised in that: the mesenchymal stem cell is prepared by following methods
Arrive: then the separation and Extraction mescenchymal stem cell from marrow is obtained through originally culture and secondary culture.
3. application according to claim 2, it is characterised in that: the extracting method of the excretion body are as follows: cultivate enough bones
Bone marrow-drived mesenchymal stem, the 48h before extracting stem cell excretion body clean cell with PBS, replace serum free medium, continue to train
Support 48h；
Serum free medium is collected, 300 × g is centrifuged 10min, takes supernatant；2000 × g is centrifuged 10min, takes supernatant；10000×g
It is centrifuged 30min, takes supernatant；100000 × g is centrifuged 70min, takes precipitating；Precipitating is resuspended in PBS, and 100000 × g is centrifuged 70min, receives
Collect and precipitates up to the excretion body.
4. application according to claim 1, it is characterised in that: the concentration of the excretion body in the drug is 1.3 × 109~
3.5×109Grain/milliliter.
5. application according to claim 1, it is characterised in that: the drug also includes pharmaceutically acceptable carrier.
6. application according to claim 1, it is characterised in that: the dosage form of the drug is injection or lyophilized preparation.
7. application according to claim 1, it is characterised in that: the drug is applied to people by way of venoclysis
Body.
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| CN107201361A (en) * | 2017-05-11 | 2017-09-26 | 广州华银医学检验中心有限公司 | A kind of extracting method of excretion body total serum IgE |
| CN110664843B (en) * | 2017-12-11 | 2022-07-01 | 浙江大学 | Application of NK cell exosome hsa-miR-9550 in antibiosis |
| CN109097328A (en) * | 2018-08-24 | 2018-12-28 | 深圳市浊安认证生物技术有限公司 | One species specific mescenchymal stem cell excretion body extracting method |
| CN109432130A (en) * | 2018-12-20 | 2019-03-08 | 中科广聚(北京)生物医学技术中心有限公司 | Application of the mescenchymal stem cell excretion body in the drug that preparation prevents and treats induced lung injury |
| CN111135190A (en) * | 2020-01-18 | 2020-05-12 | 广东医科大学 | Gingiva mesenchymal stem cell exosome preparation and application thereof |
| CN114848675A (en) * | 2022-06-24 | 2022-08-05 | 南开大学 | Application of PGE2 energized mesenchymal stem cell preparation in preparation of medicine for treating lung injury |
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