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Method for using lactoferrin for treatment or prevention of helicobacter pylori infection

A technology of Helicobacter pylori and lactoferrin, applied in peptide/protein components, antibacterial drugs, pharmaceutical formulations, etc., can solve the problems of long treatment course, inconvenient taking, lack of methods for Helicobacter pylori, etc.

Inactive Publication Date: 2016-03-30
SHANGHAI GENON BIOENG +1
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0004] In addition, there are treatment methods that use traditional Chinese medicine and a combination of Chinese and Western medicine. However, due to the complex components of Chinese medicine decoction, long treatment period, inconvenience in taking, and higher cost than Western medicine, further research is needed in terms of simplifying prescriptions and extracting active ingredients.
[0005] In summary, there is still a lack of satisfactory and safe methods for preventing and treating Helicobacter pylori in this field. Therefore, there is an urgent need in this field to develop new drugs that can effectively inhibit Helicobacter pylori infection.

Method used

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  • Method for using lactoferrin for treatment or prevention of helicobacter pylori infection
  • Method for using lactoferrin for treatment or prevention of helicobacter pylori infection
  • Method for using lactoferrin for treatment or prevention of helicobacter pylori infection

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Experimental program
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Effect test

Embodiment 1

[0152] Embodiment 1 Helicobacter pylori cultivation and cryopreservation

[0153] Helicobacter pylori (Hp) culture: Weigh 3.9g of Columbia agar, dissolve in deionized water, dilute to 90ml, autoclave (121°C, 30min), add 10ml heat inactivation after cooling to 50-60°C Fetal bovine serum (56°C, 30min), Amphotericin B 200μl (5.0mg / ml), Vancomycin 100μl (10.0mg / ml), Polymyxin B 10μl (3.8mg / ml), TMP (5.0mg / ml ) 100 μl, mixed evenly and poured into a sterile culture dish to make a solid culture plate of Hp. The frozen standard HpSS1 strain (ATCC43504) was thawed on ice, inoculated on the Hp solid culture plate, and the culture dish was placed in an anaerobic culture box, and the microaerophilic (5% O 2 , 10% CO 2 and 85%N 2 ), 90% humidity, and cultured at 37° C. for 3 to 5 days. The well-growing Hp was transferred to a new medium for subculture, and cultured under the same conditions for 3 to 5 days.

[0154] Cryopreservation of Hp: The cryopreservation solution is 3.8% brain ...

Embodiment 2

[0155] The identification of embodiment 2 Helicobacter pylori

[0156] Identification of Hp: identification by two methods

[0157] (1) Catalase experiment: Hp is a bacterium with catalase, which can catalyze hydrogen peroxide into water and oxygen, and bubbles appear. Catalyst test shows, the bacterium that is cultivated can make 3% hydrogen peroxide solution produce a large amount of bubbles (specifically as figure 1 shown).

[0158] (2) Rapid urease test:

[0159] Helicobacter pylori can produce urease and divide urea to form ammonia gas, making the medium alkaline. Pick the culture of bacteria to be tested for 18 to 24 hours and inoculate it into gastric Helicobacter pylori (HP) test paper (Guangzhou Beisiqi Diagnostic Reagent Co., Ltd.) and continue to culture for 2 to 4 hours. The result is urease positive (specifically as figure 2 shown).

Embodiment 3

[0160] Embodiment 3. Plate culture method detects the effect of rhLF inhibiting Hp bacteria

[0161]Take 6 sterile test tubes, add 6ml of 0.22μm filter-sterilized rhLF (15mg / ml) to the first tube, add 3ml of sterile water to the remaining tubes, and make 5 dilutions with the 2-fold ratio dilution method: 15, 7.5, 3.75, 1.875, 0.9375 mg / ml. The last tube without drug was used as negative control.

[0162] Take 6 sterile Erlenmeyer flasks, add 24.5ml / bottle of sterile Columbia culture, add 2.5ml / bottle of heat-inactivated fetal bovine serum, 3ml / bottle of rhLF and negative For the control, mix well and pour into a sterile Petri dish. In each group of plates, the concentration of rhLF was as follows: 1.5, 0.75, 0.375, 0.1875, 0.09375, 0 mg / ml. Plate No. 7 is a positive control with no bacteria and drugs.

[0163] Detect the OD600 value of Hp bacteria by spectrophotometer method, adjust the bacterial concentration to 1×108cfu / ml, take 100 μl of Hp bacteria suspension on the so...

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Abstract

The present invention provides a method for using lactoferrin for treatment or prevention of helicobacter pylori infection, and in particular, the present invention provides application of the lactoferrin in preparation of a composition for inhibiting helicobacter pylori and a composition for preventing or treating helicobacter-pylori-associated diseases. Tests show that recombinant lactoferrin can effectively inhibit the helicobacter pylori, and has a significant inhibiting effect on helicobacter pylori (Hp) virulence factor cagA, vacA and ure gene expression. In addition, when a lactoferrin preparation is compatibly used for a patient, the efficacy of conventional triple or quadruple therapy can be strengthened, so that the lactoferrin has a broad business prospect.

Description

technical field [0001] The invention belongs to the field of biotechnology, in particular, the invention relates to a method for using lactoferrin to treat or prevent Helicobacter pylori infection and related digestive tract diseases. Background technique [0002] Helicobacter pylori (H. pylori, Hp) is a Gram-negative microaerophilic, curved rod-shaped bacteria. It can produce a large amount of urease, which decomposes urea in the stomach to generate ammonia, which changes the physiological conditions of the stomach, triggers a local inflammatory response in the gastric mucosa, destroys the normal gastric mucosal barrier, and induces gastroduodenal ulcers. It has been shown that Helicobacter pylori infection is one of the important causes of peptic ulcer. More than 80% of patients with gastric ulcer and 95% to 100% of patients with duodenal ulcer can be positive for Hp. When Hp is successfully eradicated, the recurrence rate of peptic ulcer can be reduced to 1% to 3%. [...

Claims

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Application Information

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IPC IPC(8): A61K38/40A61K45/06A61P31/04A61P1/04A61P1/00
Inventor 成国祥马岚青刘思国邹成钢陈建泉缪应雷张爱民罗娟袁渝萍刘雪芳
Owner SHANGHAI GENON BIOENG
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