A method for screening probiotics that achieve antidepressant function through 5-hydroxytryptophan-related pathway based on cellular level

A hydroxytryptophan, cell-level technology, applied in biochemical equipment and methods, biological testing, microbial determination/inspection, etc., can solve problems such as unexplainable, high cost, and long time-consuming

Active Publication Date: 2022-02-01
JIANGNAN UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, using animals as a screening model for antidepressant probiotics is time-consuming and costly, and the specific mechanism of its action cannot be explained
Cell models have the advantages of short time-consuming, low cost, and high throughput, but there is no cell-based screening model for antidepressant probiotics

Method used

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  • A method for screening probiotics that achieve antidepressant function through 5-hydroxytryptophan-related pathway based on cellular level
  • A method for screening probiotics that achieve antidepressant function through 5-hydroxytryptophan-related pathway based on cellular level
  • A method for screening probiotics that achieve antidepressant function through 5-hydroxytryptophan-related pathway based on cellular level

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0033] Example 1: Using RIN14B cells to screen probiotics with antidepressant potential through 5-hydroxytryptophan-related pathways

[0034] Cell culture:

[0035]RIN14B (ATCC CRL-2059) cells belong to the rat insulinoma cell line, and the medium is RPMI-1640 supplemented with 1% double antibody (100U / mL penicillin and 100μg / mL streptomycin) and 10% fetal bovine serum (FBS) Complete medium (or DMEM medium). Cells were incubated at 37°C with 5% CO 2 Culture in a closed incubator, and replace the culture medium every 2-3 days according to the growth status of the cells. During passage and cryopreservation, cells were digested with 0.25% trypsin digestion solution. The digestion time is 1-2min. The number of cell passages used in this example is 10-20 passages.

[0036] Probiotic culture:

[0037] The strains used are listed in Table 1. After the strain is activated, inoculate 1% inoculum in MRS liquid medium, culture at 37°C for 20 hours, centrifuge at 3000 rpm for 10 mi...

Embodiment 2

[0052] Embodiment 2: the animal experiment verification of the effective bacterial strain of cell screening

[0053] 1. Establishment and treatment of chronic stress and depression mouse model

[0054] Seventy-eight 6-week-old male C57BL / 6J mice were randomly divided into four groups according to body weight after acclimatization for one week: control group, model group, fluoxetine intervention group, and probiotics intervention group (respectively gavage the 10 strains of probiotics), each group contained 6 mice. Animal grouping and treatment methods are shown in Table 2.

[0055] Table 2 Grouping and processing methods of animal experiments

[0056]

[0057]

[0058] Chronic unpredictable stress depression mouse model: 1-2 kinds of stimuli are randomly used every day, and the time of stimulation is randomly determined every day to avoid circadian rhythm. No more than three times of each method for a period of five weeks. Stimulus factors include: (1) fasting for 24...

Embodiment 3

[0072] Correlation Analysis of Example 3 Cell Screening Results and Animal Experiment Results:

[0073] In order to further verify the validity of the cell screening model, Pearson correlation analysis was performed on the results of cell experiments (5-HTP and TPH1) and each detection index of animal experiments. The result is as Figure 8 As shown, where blue represents positive correlation, red represents negative correlation, and the size of the dot area reflects the size of the correlation coefficient. It can be seen that the expression of TPH1 mRNA and the secretion of 5-HTP in RIN14B cells are positively correlated with the behavioral performance (reduction of depressive symptoms) and BDNF level in the prefrontal cortex in animal experiments, and negatively correlated with HPA hyperfunction. Among them, the amount of 5-HTP secreted by RIN14B cells was significantly positively correlated with the sugar preference of mice, the shortening of platform jumping latency, and ...

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Abstract

The invention discloses a method for screening probiotics that realize antidepressant function through 5-hydroxytryptophan-related pathways based on cell level. Using RIN14B cells as a screening model, through the co-incubation of probiotics and cells, the secretion of 5-hydroxytryptophan in the cell supernatant and the mRNA level of tryptophan hydroxylase 1 in the cells were detected to determine the ability to pass 5-hydroxytryptophan Hydroxytryptophan probiotics exert antidepressant effects. The effective bacterial strain screened by the present invention can alleviate the depression-like behavior of depressed mice, improve the levels of neurotransmitters and neurotrophic factors in the brain, and reduce the hyperfunction of the hypothalamus-pituitary-adrenal axis caused by stress, and the results of cell model screening are consistent with those of animals. The experimental results have a significant correlation. The antidepressant potential of different bacterial strains can be measured in batches in a short time by using the invention, and a more rapid and efficient method is provided for the screening of probiotics with antidepressant function.

Description

technical field [0001] The invention relates to the technical field of probiotic screening, in particular to a cell-based screening method for probiotics that achieve antidepressant functions through 5-hydroxytryptophan-related pathways. Background technique [0002] Depression, also known as depressive disorder, is characterized by significant and persistent depression as the main clinical feature. Currently, more than 350 million people worldwide suffer from depression, and it is estimated that by 2030, depression will rank first in the global disease burden. A large number of studies have shown that the occurrence of depression is related to the imbalance of monoamine neurotransmitters in the brain, mainly manifested as a decrease in the concentration of 5-hydroxytryptamine (5-HT) in the synaptic gap of the brain. The current treatment for depression mainly focuses on increasing the level of 5-HT in the brain, such as selective serotonin reuptake inhibitors (SSRIs), the ...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C12Q1/689C12Q1/04G01N33/68
CPCG01N33/6812C12Q1/689C12Q2600/158
Inventor 王刚陈卫田培郡赵建新张灏
Owner JIANGNAN UNIV
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