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Application of growth factor gene drug in preventing and treating stress-related gastrointestinal injury

A stress and drug technology, applied in gene therapy, microorganism-based methods, microorganisms, etc., can solve problems such as difficult control and maintenance of drug effect concentration, inability to prevent and control gastrointestinal stress injury, and reduced absorption function.

Active Publication Date: 2012-12-12
苏州厚朴惠康生物医药科技有限公司
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0003] The current treatment methods for gastrointestinal mucosal injury mainly include antibiotics, probiotics, parenteral nutrition, and traditional Chinese medicine. The barrier damage will greatly reduce the absorption function, and it is difficult to control and maintain the effective concentration of the drug; it makes the repair of gastrointestinal mucosal damage unsatisfactory
The prevention and treatment of gastrointestinal stress injury in the environment of hypoxia and high altitude is even weaker.

Method used

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  • Application of growth factor gene drug in preventing and treating stress-related gastrointestinal injury
  • Application of growth factor gene drug in preventing and treating stress-related gastrointestinal injury
  • Application of growth factor gene drug in preventing and treating stress-related gastrointestinal injury

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0086] Example 1. Construction and preparation of recombinant attenuated Salmonella SPH

[0087] 1. Cloning of HIF1α cDNA

[0088] Human hypoxia inducible factor-1α, as reported in literature (Miyazawa K, et al BBRC, 1989, 163:967-973; Nakamura T, et al. Progress in Growth Factor Research 1991, 3:67-85), Primers were designed based on HIF1α) cDNA sequence, Nhe 1 restriction site was introduced into the forward primer, and Mlu I restriction site was introduced into the reverse primer. The oligonucleotide sequence of the forward primer is 5'-cat gctagc cac cga ttc accatg gag ggc-3' (SEQ ID NO: 1), and the oligonucleotide sequence of the reverse primer is 5'-gtc acgcgt tca gtt aac ttg atc caa agc tctg-3' (SEQ ID NO: 2). In this paper, unless otherwise specified, the primers mentioned are all synthesized by Takara Company.

[0089] Use a 25ml cell culture flask 1640 medium (Gibco) in a trigas incubator at 37°C 5% CO 2 Under the conditions, the Hela cells were induced to be c...

Embodiment 2

[0103] Example 2. Recombinant attenuated Salmonella carrying KGF gene (Ty21a-pIRES-KGF (TPK)) construction and preparation

[0104] 1. Cloning of KGF cDNA

[0105]Primers were designed according to the human KGF cDNA sequence reported in the literature (Rubin JS, et al Proc Natl Acad Sei USA1989.86: 302-306; Finch PW, et al Science 1989, 245 (49 19): 752-755), in the forward direction The Xbal restriction site was introduced into the primer, and the NotI restriction site was introduced into the reverse primer. The oligonucleotide sequence of the forward primer is 5'-TATTCTCAGAATGAGCTATGATTACATGGAAG-3' (SEQ ID NO: 4), and the oligonucleotide sequence of the reverse primer is 5'-GACGCGGCCGCTTAAGTGATTGCCATAGGCAG-3' (SEQ ID NO: 5). The human keratinocyte growth factor (KGF) gene was cloned from a human placenta cDNA library (purchased from Clontech). Reaction system: template cDNA 1 μl, KGF upstream primer (20 μM) and downstream primer (20 μM) 1 μl each, dNTP 3 μl, 10×PCR ...

Embodiment 3

[0118] Example 3. Recombinant attenuated Salmonella carrying HIF-1α gene and KGF gene Construction and Preparation of (Ty21a-pIRES-HIF1α-KGF(TPHK))

[0119] 1. Construction of double-gene plasmid carrying human KGF and HIF-1α

[0120] The PCR product of the KGF gene in Example 2 was gel-cut and recovered, ligated with the Xba I and Not I (both purchased from Takara) double digestion products of the pIRES-HIF-1α vector, and the resulting recombinant plasmid was named pIRES-HIF- 1α-KGF, its plasmid map is as follows Figure 17 shown.

[0121] Determine the human HIF1α cDNA sequence (Shanghai Sangon Bioengineering Technology Service Company). The sequence of the obtained pIRES-HIF-1α-KGF is shown in SEQ ID NO:8.

[0122] 3. Screening and preparation of an attenuated Salmonella strain (Ty21a-pIRES-HIF1α-KGF (TPHK)) containing the eukaryotic expression plasmid pIRES-HIF-1α-KGF

[0123] 3.1. Electroporation transformation

[0124] Inoculate 50 μL of the attenuated Salmo...

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Abstract

The invention relates to a cell growth factor gene drug for preventing and / or treating acute stress-related gastrointestinal injury, and use of HIF-1 alpha and KGF genes in preparing drugs for preventing and / or treating acute stress-related gastrointestinal injury diseases. The invention also relates to a pharmaceutical composition comprising HIF-1 alpha and KGF. The growth factor gene drug provided by the invention can beneficially prevent and / or treat acute stress-related gastrointestinal injury diseases.

Description

technical field [0001] The present invention relates to the application of cell growth factor gene medicine in the prevention and treatment of stress gastrointestinal injury, in particular to hypoxia-inducible factor-1α (hypoxia-inducible factor-1α, HIF-1α) and keratinocyte growth factor (keratinocyte growth factor, The combination of KGF) in the prevention and / or treatment of stress-induced gastrointestinal injury diseases, more particularly relates to its application in the prevention and / or treatment of acute stress-induced gastrointestinal injury diseases. Background technique [0002] The gastrointestinal mucosa is an important place for the digestion and absorption of nutrients. However, a variety of stress factors, including pathogenic microorganisms, physical factors, chemical factors, mechanical factors, etc. damage. At the same time, the gastrointestinal mucosal barrier, as one of the central organs of the stress response, is involved in the occurrence, developmen...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): A61K48/00A61K38/18A61K9/48A61P1/04C12N15/79C12N1/21C12R1/42C12R1/01C12R1/19
Inventor 哈小琴杨志华彭俊华邓芝云董菊子赵勇张媛媛
Owner 苏州厚朴惠康生物医药科技有限公司
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