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Use of decidual NK cells and cell subgroups in the preparation of drugs for the treatment of infertility-related diseases

A technology of NK cells and cell subgroups, which is applied in the field of biomedicine, can solve problems such as not disclosed or prompted, and achieve the effect of promoting VEGF expression, promoting cell vitality, and reducing endometrial cell damage

Active Publication Date: 2021-08-27
PHARCHOICE THERAPEUTICS INC
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

The prior art does not disclose or suggest a technical solution for preparing products from decidual NK cell subsets expressing specific markers for disease treatment

Method used

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  • Use of decidual NK cells and cell subgroups in the preparation of drugs for the treatment of infertility-related diseases
  • Use of decidual NK cells and cell subgroups in the preparation of drugs for the treatment of infertility-related diseases
  • Use of decidual NK cells and cell subgroups in the preparation of drugs for the treatment of infertility-related diseases

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0026] Example 1 Screening of uterine decidua NK cell candidate markers

[0027] Statistical calculations in this example and the examples described later calculate the p-values ​​according to different statistical modes of the software SPSS version 22.0 for two-group comparison, multi-group comparison, and rate comparison. A P value of less than 0.05 was considered statistically significant.

[0028] Firstly, NK cells were isolated from the decidual tissues of 10 cases of healthy non-medical termination of early pregnancy (normal group) and 5 cases of spontaneous abortion early pregnancy decidual tissues (abortion group). The separation and flow cytometry of NK cells can be referred to [ Fu B, et al. Immunity, 2017, 47(6):1100-1113.e6.]. An example is as follows: Lymphocytes were obtained by digesting with 1 mg / mL collagenase IV (Sigma-Aldrich) and 0.01 mg / mL DNase I (Shanghai Sangon) for 1 h, and then centrifuging with Percoll (GE Healthcare) density gradient. Culture dish...

Embodiment 2

[0029] Example 2 Preparation of uterine decidua NK cells and decidua NK cell subsets

[0030] Utilize healthy non-medical reasons to terminate early pregnancy decidual tissue to prepare decidual NK cells, implement according to the NK cell separation method described in Example 1, briefly as follows: After 1 mg / mL collagenase IV (Sigma-Aldrich) and 0.01 mg / mL Lymphocytes were obtained by digesting with DNase Ⅰ (Shanghai Sangon) for 1 h, and then centrifuging with Percoll (GE Healthcare) density gradient. Culture dishes were cultured at 37°C for 2 hours to remove stromal cells and macrophages, and then NK cells were separated by flow cytometry. The resulting phenotype is CD56 bright CD16 - CD49a + decidua NK cells. Further use of antibody magnetic beads to sort CD56 bright CD16 - C D49a + CD39 + Decidual NK cell population, CD56 bright CD16 - CD49a + CD27 + Decidual NK cell population, CD56 bright CD16 - CD49a + CD160 + Decidual NK cell population, CD56 bright...

Embodiment 3

[0031] Example 3 Decidual NK cells and decidual NK cell subsets enhance endometrial cell viability, reduce endometrial cell damage, and increase VEGF expression

[0032] After culturing non-pathological endometrial stromal cells (stroma cells) for 24 hours, each group of NK cells described in Example 2 was added, and the ratios of stromal cells: NK cells were 10:1 and 5:1, respectively. The control group used the control NK cells described in Example 2. NK cells were cleared 48 hours after treatment, and stromal cells and media were sampled for analysis.

[0033] Cell viability of stromal cells was detected by the PrestoBlue method (Thermo Fisher Scientific), measured 48 hours after treatment, and values ​​are expressed as mean % normalized to control (Table 1 and Table 2).

[0034] Table 1 Relative cell viability of stromal cells under 10:1 culture condition

[0035] average value SD p-value Blank (cultured alone, no NK cells) 100 6.72 control c...

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Abstract

The invention provides the use of decidual NK cells and cell subgroups in the preparation of drugs for treating infertility-related diseases. It has been confirmed by experiments that the decidual NK cells and their cell subsets can promote endometrial thickness increase, enhance endometrial cell viability, reduce endometrial cell damage, promote VEGF expression, maintain endometrial stromal cell stemness and stimulate proliferation. The method of treating endometrial growth disorder disease can increase the pregnancy success rate of endometrial injury model mice from 30% to 60-70%; The treatment of diseases related to maternal-fetal immune tolerance disorder at the level of lymphocytes provides a new way for the treatment of infertility.

Description

technical field [0001] The invention belongs to the field of biomedicine, and relates to the use of decidual NK cells and cell subgroups in preparing medicines for treating infertility-related diseases, a method for separating and culturing cells, and a pharmaceutical composition containing the same. Background technique [0002] The endometrial injury is mainly the injury of the endometrial basal layer, which is mainly related to curettage during pregnancy. Compared with the normal endometrium, the endometrial basal layer during pregnancy is loose and more susceptible to damage. Based on my country's national conditions and the increasing rate of induced abortion, the occurrence of endometrial damage cannot be ignored. The endometrial basal layer is damaged, which may lead to damage or loss of endometrial stem cells; at the same time, local infection and aseptic inflammation of the injured endometrium can destroy the niche microenvironment of stem cells, resulting in regene...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C12N5/0783A61K35/17A61P15/08
CPCA61K35/17A61P15/08C12N5/0646C12N2509/00
Inventor 胡适
Owner PHARCHOICE THERAPEUTICS INC
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