CN106581666A - Application of PEDF in preparing medicine for diabetes bone tissue complication - Google Patents
Application of PEDF in preparing medicine for diabetes bone tissue complication Download PDFInfo
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- CN106581666A CN106581666A CN201611036009.3A CN201611036009A CN106581666A CN 106581666 A CN106581666 A CN 106581666A CN 201611036009 A CN201611036009 A CN 201611036009A CN 106581666 A CN106581666 A CN 106581666A
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- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
- A61K38/00—Medicinal preparations containing peptides
- A61K38/16—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof
- A61K38/55—Protease inhibitors
- A61K38/57—Protease inhibitors from animals; from humans
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Abstract
The invention discloses application of PEDF in preparing medicine for diabetes bone tissue complication, and most diabetes patients have bone tissue complication, including osteoporosis, bone healing difficulty, gangrene and the like,. When PEDF expression levels of an osteoblast and osteogenic capability of a diabetes patient and a non-diabetes patient are compared, it is found that PEDF expression level of the non-diabetes patient is higher, and the osteogenic differentiation ability, the osteogenic efficiency and the calcium deposition level of the non-diabetes patient are higher than those of the diabetes patient (p<0.01). Exogenetic PEDF can up-regulate expression of osteogenic differentiation related genes OCN and Runx2. Compared with VEGF expression levels of the osteoblast of the two groups of the patients, it is found that the overall VEGF level of the non-diabetes patient is higher than that of a diabetes patient, and the exogenetic PEDF can up regulate expression of VEGF. PEDF can improve osteogenic differentiation ability of the osteoblast and calcium deposition level. PEDF can be applied to treating bone tissue complication caused by diabetes.
Description
Technical field
The invention belongs to biomedicine field, and in particular to PEDF is in treatment diabetes osseous tissue complication medicine is prepared
Application.
Background technology
Diabetes not only cause including sugar, and fat, the exception of protein three major nutrient further relate to electrolyte, bone ore deposit
The metabolism disorders such as material.Therefore the complication that diabetes cause is the greatest problem for facing at present, and for example patient wound is long-term not
Healing, etc. symptom.Osseous tissue is also to be affected to organize than larger one by diabetic complication, there are some researches show, glycosuria
Patient easily causes osteoporosis due to the decline of doped calcium concentration, is difficult healing in addition after patient's fracture.It has been reported that sugared
Urine patient is low compared with normal person in budding bone formation and cumulant, and obstacle occurs in terms of bone resorption again afterwards, easily goes out
Existing osteoporotic symptom.In the diabetes study model of animal, it has been found that doped calcium level, bone mineral content and bone
The expression of calcium element is decreased obviously.
Gangrenous problem is also the complication for making diabeticss overwhelmed with sorrow, result in the probability calibration of diabetes patient's amputation
The probability of often patient's amputation is high.The principal element for causing gangrenous problem is exactly that blood glucose is high, will cause machine in the case of blood glucose height
The vascular lesion of body, neuropathy, the problems such as leukocyte changes.Osseous tissue is euangiotic tissue, and VEGF is blood vessel endothelium
Angiogenesis are had facilitation by somatomedin.Prevent the activity of VEGF-A with antibody VEGFR1 and VEGFR2, can significantly reduce
The formation of osseous tissue, blood vessel number are substantially reduced.The increase of blood vessel number can promote the maturation of osteoprogenitor cells, increase osseous tissue
Formation.It is likely to accelerate osteogenetic process by exciting blood vessel signal simultaneously.
Therefore, by raising diabetic osseous tissue vegf expression level, and promotion diabetic bone doped calcium
By effective diabetes-alleviating complication.
The content of the invention
The purpose of the present invention is to cause osseous tissue to go wrong the invention aims to solve diabetic complication, is wrapped
Osteoporosises, bone injury healing difficulty, gangrene etc. are included, and a kind of rise osteoblast vegf expression and OCN and Runx2 genes are provided
Expression, so as to improving diabetic osteoporosises, promoting bone injury healing and alleviating the medicine PEDF of gangrene.
PEDF is promoting the purposes of osteoblastic vegf expression.
PEDF is improving the purposes of osteoblast OCN and Runx2 gene expression.
Applications of the PEDF in diabetes osseous tissue complication medicine is prepared.
The gangrene that described osseous tissue complication is osteoporosises, bone injury healing is difficult or osseous tissue pathological changes cause.
The invention provides applications of the PEDF in treatment diabetes osseous tissue complication medicine is prepared, diabetic complication
Osseous tissue is caused to go wrong, including osteoporosises, knitting is difficult, gangrene etc..Our experiments show that, PEDF can affect mesenchyme
The efficiency and doped calcium level of stem cell Osteoblast Differentiation.The PEDF of external source can accelerate the Osteoblast Differentiation process of mesenchymal cell,
Improve doped calcium level.Osteoblast derives from mescenchymal stem cell, compares diabetes patient and non-diabetic human osteoblast cell
Find after the expression and Osteoblast Differentiation ability of PEDF, the PEDF expressions of non-diabetic people are high, and Osteoblast Differentiation energy
Power, efficiency and doped calcium level it is all high than diabetes patient（p<0.01）.External source PEDF can raise diabetes patient's Osteoblast Differentiation
The expression of related gene OCN, Runx2.The osteoblastic vegf expression level discovery of two groups of patients of comparison, non-diabetic people's entirety
VEGF levels it is higher than the VEGF levels of diabetes patient, the PEDF of external source can raise the expression of VEGF.PEDF can improve sugar
Urine patient's osteoblast differentiation ability, doped calcium level.May apply to treat the osseous tissue correlation that diabetic complication causes
Disease.
Description of the drawings
Fig. 1 separates osteoblast cultivation results；
Fig. 2 PEDF raise the expression of results of osteoblast VEGF；
Fig. 3 PEDF raise OCN and Runx2 gene expression results；
Fig. 4 PEDF promote diabetic osteoblast doped calcium result.
Specific embodiment
The osteoblastic separation of embodiment 1 and culture
The bone garbage sample of sufficient amputation patient is collected, is divided into diabetes patient's group and non-diabetic people's group.Tissue samples are by Binzhou
The uncommon medical college in state university Hull is provided.By the tissue samples collected with PBS rinses 3 times after, remove the knot on osseous tissue surface
Form tissue and fatty tissue, and with ethanol postincubation cell 15s（Remove the fibroblast on osseous tissue surface）.With concentration it is
The collagenase digesting osseous tissue of 4mg/ml, collects a Digestive system supernatant per 45min, and 1000rpm centrifugation 5min stay precipitation, even
Continuous operation three times, after the 4th peptic cell 45min, is collected by centrifugation osseous tissue and unicellular precipitation.They are uniformly taped against
Cultivate in 100mm Tissue Culture Dishs.Culture fluid composition is that a-MEM, 10%FBS, 1% are dual anti-, 37 °, 5% CO27 are cultivated in incubator
After it, osseous tissue is discarded, change fresh medium.Well-grown, such as Fig. 1 are observed under osteoblast inverted microscope after culture
It is shown.
2 PEDF of embodiment raises the level of diabetes patient osteoblast VEGF
By the condition of culture of embodiment 1, add the PEDF of 250ng/ml in skeletonization culture fluid, using conventional qPCR technology for detection
The level of VEGF, qPCR testing conditions be 95 DEG C of 5min, 40 ×（95 DEG C of 3s, 60 DEG C of 30s）, solubility curve is 95 DEG C of 15s
（100%）；65 DEG C of 1min, 95 DEG C of 15s（100%）,+0.3 DEG C/circle, b-actin is used as internal reference.As a result show, 250ng/
Ml PEDF process cell vegf expression rise after 7 days.As shown in Figure 2.
3 PEDF of embodiment raises the expression of OCN and Runx2 genes
OCN and Runx2 are osteoblastic marker gene.By the condition of culture of embodiment 1, add in skeletonization culture fluid
The PEDF of 250ng/ml, using conventional qPCR technology for detection OCN and the expression of Runx2 genes, qPCR testing conditions are 95
DEG C 5min, 40 ×（95 DEG C of 3s, 60 DEG C of 30s）, solubility curve is 95 DEG C of 15s（100%）；65 DEG C of 1min, 95 DEG C of 15s
（100%）,+0.3 DEG C/circle, b-actin is used as internal reference.As a result show, 250ng/ml PEDF process cell after 7 days OCN and
Runx2 gene expressions are raised.As shown in Figure 3.
4 PEDF of embodiment promotes the osteoblastic doped calcium of diabetic
Osteoblast is inoculated in 6 orifice plates, with Osteoblast Differentiation liquid cultured cells, liquid is changed once per two days.Osteoblast Differentiation liquid into
Be divided into the DMEM in high glucose culture medium of 10%FBS, the ascorbic acid of 50ug/ul, the β phosphoglycerol salt of 10mM, 10-6 dexamethasone with
And 1% it is dual anti-.In experimental group, the pedf protein of addition 250ng/ml is in skeletonization culture fluid.Collection of cellular samples after 14 days
Alizarin red staining analysis cell calcium deposition level is carried out, doped calcium level is higher, and alizarin red coloring is deeper.As a result show,
The PEDF of 250ng/ml can dramatically increase the osteoblastic doped calcium level of diabetic.As shown in Figure 4.
5 PEDF of embodiment improves diabetic model rats osseous tissue complication
Using the PEDF intravenous injection diabetic model rats of 0.25mg/kg body weight, once a day.Examined at 7,14,28 days respectively
Survey the expression of rat bone doped calcium and osseous tissue VEGF.As a result show, after 14 days, compared with matched group, bone calcium sinks PEDF
Product and osseous tissue vegf expression level are dramatically increased.
Claims (4)
1.PEDF is promoting the purposes of osteoblastic vegf expression.
2.PEDF is improving the purposes of osteoblast OCN and Runx2 gene expression.
Applications of the 3.PEDF in diabetes osseous tissue complication medicine is prepared.
4. application according to claim 3, it is characterised in that：Described osseous tissue complication is osteoporosises, bone injury medicine
Close the gangrene that difficult or osseous tissue pathological changes cause.
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| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN201611036009.3A CN106581666A (en) | 2016-11-23 | 2016-11-23 | Application of PEDF in preparing medicine for diabetes bone tissue complication |
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| Application Number | Priority Date | Filing Date | Title |
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| CN201611036009.3A CN106581666A (en) | 2016-11-23 | 2016-11-23 | Application of PEDF in preparing medicine for diabetes bone tissue complication |
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| CN106581666A true CN106581666A (en) | 2017-04-26 |
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Citations (4)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN1856326A (en) * | 2003-09-23 | 2006-11-01 | 诺瓦提斯公司 | Combinations of a VEGF receptor inhibitor with other therapeutic agents |
| CN101616693A (en) * | 2006-11-24 | 2009-12-30 | 泰根尼克斯股份有限公司 | Be used to identify that chondrocyte phenotypic stability and screening cartilage form the marker gene of factor of influence |
| CN102895652A (en) * | 2011-07-27 | 2013-01-30 | 东北师范大学 | Method for promoting osteoblast differentiation by using Runx2 and Osterix and application thereof |
| CN104903344A (en) * | 2012-09-20 | 2015-09-09 | 财团法人台湾基督长老教会马偕纪念社会事业基金会马偕纪念医院 | Use of pedf-derived polypeptides for treating osteoarthritis |
-
2016
- 2016-11-23 CN CN201611036009.3A patent/CN106581666A/en active Pending
Patent Citations (4)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN1856326A (en) * | 2003-09-23 | 2006-11-01 | 诺瓦提斯公司 | Combinations of a VEGF receptor inhibitor with other therapeutic agents |
| CN101616693A (en) * | 2006-11-24 | 2009-12-30 | 泰根尼克斯股份有限公司 | Be used to identify that chondrocyte phenotypic stability and screening cartilage form the marker gene of factor of influence |
| CN102895652A (en) * | 2011-07-27 | 2013-01-30 | 东北师范大学 | Method for promoting osteoblast differentiation by using Runx2 and Osterix and application thereof |
| CN104903344A (en) * | 2012-09-20 | 2015-09-09 | 财团法人台湾基督长老教会马偕纪念社会事业基金会马偕纪念医院 | Use of pedf-derived polypeptides for treating osteoarthritis |
Non-Patent Citations (1)
| Title |
|---|
| 王海军等: "糖尿病人的成骨细胞中分化以及成骨相关基因表达方面的差异", 《中国生物工程学会2014年学术年会暨全国生物技术大会论文集》 * |
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