阅读说明：本技术 Pgam5作为少弱精症诊断标志物及治疗靶点的应用 (Application of PGAM5 as diagnosis marker and treatment target of oligoasthenospermia ) 是由 黄宁 董锡阳 张亚卓 张曼玉 孙胜楠 于 2020-06-23 设计创作，主要内容包括：本发明属于少弱精诊断技术领域,具体涉及PGAM5作为少弱精症诊断标志物及治疗靶点的应用。本发明基于蛋白质组学方法,对正常和少弱精患者样本中的蛋白数据进行分析,以BSA蛋白信号强度为标准,检测上述样本中的蛋白丰度获得正常与患病样本中的差异蛋白。本发明通过该方法检测确认PGAM5在重度少弱精样本中显著下调,作为诊断生物标志物具有良好的可信度,应用于相关检测试剂及试剂盒的开发。同时,上述研究结果有助于明确少弱精症的发病机制,有助于少弱精症靶点药物的开发,具有重要的临床意义。(The invention belongs to the technical field of oligozoospermia diagnosis, and particularly relates to application of PGAM5 as an oligozoospermia diagnosis marker and a treatment target. The invention is based on a proteomics method, analyzes protein data in normal and oligozoospermia patient samples, detects protein abundance in the samples by taking BSA protein signal intensity as a standard, and obtains differential protein in normal and diseased samples. The method provided by the invention is used for detecting and confirming that PGAM5 is obviously reduced in severe asthenospermia samples, has good reliability as a diagnosis biomarker, and is applied to development of related detection reagents and kits. Meanwhile, the research result is helpful for determining the pathogenesis of oligospermia and developing a target drug for the oligospermia, and has important clinical significance.)

The application of PGAM5 as a biomarker in preparing a product for diagnosing oligospermia, asthenospermia or severe oligospermia.

2. Use of PGAM5 as a biomarker for the manufacture of a diagnostic product for oligozoospermia, asthenospermia or severe oligozoospermia according to claim 1, wherein the diagnostic product comprises a diagnostic reagent or a diagnostic kit.

3. The use of PGAM5 as a biomarker for the preparation of diagnostic products for oligospermia, asthenospermia or severe oligospermia according to claim 2, wherein the diagnostic reagents include but are not limited to reagents for measuring the content of PGAM5 in a sample by immunoblotting, enzyme linked immunosorbent assay, chromatography or genetic engineering; the diagnostic kit comprises the reagent for detecting the content of PGAM5 in the sample.

The application of PGAM5 as a biomarker in preparing medicines for treating oligospermia, asthenospermia and severe oligospermia.

5. Use of PGAM5 as a biomarker for the manufacture of a medicament for the treatment of oligospermia, asthenospermia and severe oligospermia according to claim 4, wherein the use comprises the use of an active substance which modulates the content of PGAM5 for the manufacture of a medicament for the treatment of oligospermia, asthenospermia and severe asthenospermia.

6. The use of PGAM5 as a biomarker for the manufacture of a medicament for the treatment of oligospermia, asthenospermia and severe oligospermia according to claim 5, wherein the active substance that modulates the level of PGAM5 is an agonist of PGAM 5.

7. A kit for diagnosing oligoasthenospermia is characterized by comprising a reagent for detecting the content of PGAM5 in a sperm sample.

8. A medicament for the treatment of oligoasthenospermia, which comprises an active ingredient that modulates the level of expression of PGAM 5.

9. The medicament for treating oligoasthenospermia according to claim 8, wherein the active ingredient is a PGAM5 agonist.

10. A method for diagnosing asthenospermia, which comprises the steps of: extracting a sperm sample of an object to be detected, detecting the detection abundance of PGAM5 protein in the sperm, and judging the patient with oligozoospermia when the detection abundance is less than 0.00591.

Technical Field

The invention belongs to the field of oligozoospermia diagnostic markers, and particularly relates to application of PGAM5 as a diagnostic marker or a treatment target for oligozoospermia, asthenospermia and even severe oligozoospermia in preparation of a diagnostic reagent or a medicament for treating oligozoospermia, asthenospermia and severe oligozoospermia.

Background

The information in this background section is only for enhancement of understanding of the general background of the invention and is not necessarily to be construed as an admission or any form of suggestion that this information forms the prior art that is already known to a person of ordinary skill in the art.

According to the world health organization investigation, 15% of fertile couple have sterility problems, and infertility has become a global medical and social problem affecting human health and social development (Turner, T.T. and J.J.Lysiak, oxidative stress: a common factor in physiological stress.J. Androl,2008.29(5): p.488-98). By definition by the world health organization, asthenospermia is meant less than 32% of the sperm in the semen that move forward. Asthenospermia is one of the major causes of male infertility. Asthenospermia accounts for about 19% of male infertility, while oligospermia or teratospermia accounts for 63% of male infertility. Although many factors such as lifestyle, environmental pollution, varicocele, and infection may cause asthenospermia, which seriously affects the quality of life of people.

Asthenospermia refers to a condition in which less than 50% of the sperm in forward movement (grade a and b) or less than 25% of the sperm in grade a movement in the semen parameters, which is also called asthenospermia. The oligospermia refers to abnormal expression that the number of sperms in one milliliter of semen is less than 2000 ten thousand, and when the density of the sperms is less than or equal to 1 × 10/mL, the oligospermia is diagnosed as severe. The concept of oligospermia and asthenospermia is different, the diagnosis standard is different, but the oligospermia and asthenospermia belong to the quality problem of sperms, and the causes of the diseases are similar. At present, there is no unified understanding on the induction factors and development mechanism of oligospermia, and the factors are generally considered to be related to infection, gene mutation, vitamin deficiency and the like. Aiming at the treatment of oligospermia, no effective medicine exists, the traditional Chinese medicine for nourishing yin and tonifying kidney is mostly adopted for clinical treatment, or the traditional Chinese medicine and western medicine are combined for treatment, and the treatment mode mainly supplements vitamins and tonifies essence, qi and blood deficiency of kidney. In order to overcome the medical problem, the development of related mechanism research is beneficial to determining the pathogenesis of oligospermia and correspondingly researching and developing corresponding target drugs.

PGAM5(The protein phosphate mutase 5) is a serine/threonine protein phosphatase, activation of PGAM5 can lead to linear fragmentation of mitochondria arranged in a string, and is related to apoptosis, necrosis and autophagy of mitochondria, and PGAM5 plays a pivotal role in a pathway of cellular necrosis caused by various causes, and is involved in cellular necrosis caused by, for example, excessive increase of oxygen radicals and excessive leakage of calcium ions, but The mechanism of cellular necrosis caused by PGAM5 is currently poorly understood.

Disclosure of Invention

Based on the research background, the invention aims to screen and research related targets and pathways of oligoasthenospermia based on sperm proteomics, and provide a basis of related mechanisms for diagnosis and treatment of severe oligoasthenospermia. According to the screening result of the invention, the protein content of PGAM5 in the oligozoospermia sample is obviously reduced, namely the relative abundance of PGAM5 in healthy and oligozoospermia samples is different, indicating that PGAM5 is expected to be a diagnosis and treatment target of oligozoospermia.

Based on the research results, the invention provides the following technical scheme:

in a first aspect of the invention, the invention provides the use of PGAM5 as a biomarker in the preparation of a product for diagnosing oligospermia, asthenospermia or severe oligospermia.

The invention detects the relative abundance of protein in sperm samples of patients with normal and severe oligozoospermia by a proteomics method, wherein the expression difference of PGAM5 in the normal sample and the patient sample is obvious, the expression content of PGAM5 in the patient sample is reduced four times, and the obvious correlation between PGAM5 and the diseased condition is proved.

The diagnostic product comprises a diagnostic reagent or a diagnostic kit; the diagnostic reagent includes but is not limited to a reagent for measuring the content of PGAM5 in the sample by an immunoblotting method, an enzyme-linked immunosorbent assay method, a chromatographic analysis method or a genetic engineering method; the diagnostic kit comprises the reagent for detecting the content of PGAM5 in the sample.

In a second aspect of the invention, the invention provides the use of PGAM5 as a biomarker in the preparation of a medicament for the treatment of oligospermia, asthenospermia and severe oligospermia.

In some specific embodiments, the use comprises the use of an active substance that modulates the level of PGAM5 for the preparation of a medicament for the treatment of oligospermia, asthenospermia and severe oligospermia. Specifically, the active substance for regulating the content of the PGAM5 is an agonist of PGAM 5.

In a third aspect of the invention, a kit for diagnosing oligoasthenospermia is provided, wherein the kit comprises a reagent for detecting the content of PGAM5 in a sperm sample.

In a fourth aspect of the present invention, there is provided a medicament for treating oligoasthenospermia, which comprises an active ingredient that modulates the expression level of PGAM 5.

Preferably, the active ingredient is a PGAM5 agonist.

In a fifth aspect of the present invention, there is provided a method for diagnosing asthenospermia, comprising the steps of: extracting a sperm sample of an object to be detected, detecting the detection abundance of PGAM5 protein in the sperm, and judging the patient with oligozoospermia when the detection abundance is less than 0.00591.

The beneficial effects of one or more technical schemes are as follows:

the invention takes the sperm sample of oligoasthenospermia as a research object, detects the relative abundance of the protein in the normal sample and the patient sample, and obtains the differential protein with higher detection value by screening. Through mass spectrometry data analysis, the expression content of PGAM5 in a patient sample is remarkably reduced by 4.1 times, the ROC analysis result shows that the PGAM5 used as a diagnostic marker has a good diagnostic effect, and the false positive probability applied to individual detection is 12.7%, so that the protein is proved to be a marker with higher accuracy and diagnostic value.

Drawings

The accompanying drawings, which are incorporated in and constitute a part of this specification, are included to provide a further understanding of the invention, and are incorporated in and constitute a part of this specification, illustrate exemplary embodiments of the invention and together with the description serve to explain the invention and not to limit the invention.

FIG. 1 is a ROC plot of the relative abundance of the PGAM5 protein assay in example 1.

FIG. 2 is a graph comparing the intensity of PGAM5 protein detection in healthy and oligozoospermic samples in example 1.

Detailed Description

It is to be understood that the following detailed description is exemplary and is intended to provide further explanation of the invention as claimed. Unless defined otherwise, all technical and scientific terms used herein have the same meaning as commonly understood by one of ordinary skill in the art to which this invention belongs.

It is noted that the terminology used herein is for the purpose of describing particular embodiments only and is not intended to be limiting of exemplary embodiments according to the invention. As used herein, the singular forms "a", "an" and "the" are intended to include the plural forms as well, and it should be understood that when the terms "comprises" and/or "comprising" are used in this specification, they specify the presence of stated features, steps, operations, devices, components, and/or combinations thereof, unless the context clearly indicates otherwise.

As described in the background, the present invention addresses the deficiencies of the prior art by providing the use of PGAM5 as a diagnostic marker and therapeutic target for severe oligoasthenospermia.

In order to make the technical solutions of the present invention more clearly understood by those skilled in the art, the technical solutions of the present invention will be described in detail below with reference to specific embodiments.