Application of lncRNA-92467 in early warning or diagnosis of altitude sickness

Abstract

The invention provides an application of lncRNA-92467 as a marker in preparation of a product for early warning and / or diagnosing altitude sickness. Particularly, the expression level of lncRNA-92467 in pulmonary arteries or cells of rats in a simulated plateau environment is remarkably reduced. And the regulation of the expression level of the lncRNA-92467 is directly related to the proliferation, migration and apoptosis of the HPAECs cells.

Description

Technical Field

[0001] The present invention relates to the field of biomedicine technology, and in particular to the application of LncRNA-92467 in early warning or diagnosis of altitude sickness. Background Art

[0002] Altitude sickness refers to a special disease characterized by metabolic changes, physiological decompensation and target organ damage during life and work at high altitudes, with the body's ability to adapt to plateau hypoxia, especially decreased work ability, as the main manifestation. It is divided into acute and chronic categories. When the body cannot adapt to this acute stimulation in a short period of time, acute altitude sickness may occur, manifested as acute mountain reaction, high-altitude cerebral edema, high-altitude pulmonary edema, etc. These manifestations can occur within a few hours to 5 days after ascending to a specific altitude, seriously affecting the individual's normal quality of life and even endangering life safety. Chronic altitude sickness is a condition in which symptoms last for more than 3 months after acute altitude reaction.

[0003] LncRNA-92467 is a long non-coding RNA newly screened by whole transcriptome sequencing, with a total length of 12061bp, located on chromosome 1. At present, there is no report in the prior art on the correlation between LncRNA-92467 and altitude sickness, and the use of LncRNA-92467 for early warning and prevention of altitude sickness. In view of this, the present invention is proposed. Summary of the invention

[0004] The first object of the present invention is to provide a reagent for detecting lncRNA-92467 or the use of lncRNA-92467 as a marker in the preparation of products for early warning and / or diagnosis of altitude sickness.

[0005] The second object of the present invention is to provide a lncRNA-92467 regulator or the use of lncRNA-92467 as a target in improving altitude tolerance or in the preparation of a drug for treating and / or preventing altitude sickness.

[0006] The third object of the present invention is to provide cells, tissues or organs that regulate the expression level of lncRNA-92467.

[0007] The fourth object of the present invention is to provide a method for constructing cells, tissues or organs that regulate the expression level of lncRNA-92467.

[0008] A fifth object of the present invention is to provide a diagnostic kit.

[0009] A sixth object of the present invention is to provide a method for early warning and / or diagnosis of altitude sickness.

[0010] A seventh object of the present invention is to provide a medicine for treating and / or preventing altitude sickness.

[0011] The eighth object of the present invention is to provide a method for treating and / or preventing altitude sickness.

[0012] The ninth object of the present invention is to provide a method for altitude tolerance.

[0013] The tenth object of the present invention is to provide a disease animal model, which comprises introducing a lncRNA-92467 inhibitor into a non-human animal.

[0014] In order to achieve the above objectives, the following technical solutions are adopted:

[0015] In a first aspect, a reagent for detecting lncRNA-92467 or the use of lncRNA-92467 as a marker in the preparation of a product for early warning and / or diagnosis of altitude sickness is provided.

[0016] The ID of the lncRNA-92467 is ENSRNOT00000092467.

[0017] The lncRNA-92467 is a long non-coding RNA with a total length of 12061 bp and is located on chromosome 1.

[0018] The lncRNA-92467 is lncRNA-92467.1.

[0019] The sequence of the lncRNA-92467 is shown in SEQ ID NO: 1.

[0020] The lncRNA-92467 is lncRNA-92467 in the pulmonary artery or cells thereof.

[0021] Preferably, the cells are pulmonary artery endothelial cells.

[0022] The product includes a reagent for detecting lncRNA-92467, preferably a reagent for detecting the expression level of lncRNA-92467.

[0023] The detection of lncRNA-92467 can be carried out by polymerase chain reaction (PCR), Southern blot hybridization, Northern blot hybridization, dot hybridization, fluorescence in situ hybridization (FISH), DNA microarray, high-throughput sequencing platform, chip detection, etc., especially PCR methods, such as real-time fluorescence quantitative PCR method, constant temperature amplification technology (rolling circle nucleic acid amplification, loop-mediated isothermal amplification, chain replacement amplification, etc.).

[0024] The reagents for detecting lncRNA-92467 include but are not limited to primers, probes, etc.

[0025] Preferably, the reagents include reagents required for reverse transcription, reagents required for PCR, reagents required for sequencing, reagents required for mass spectrometry and / or buffer.

[0026] The product can be a diagnostic kit, a sequencing platform or a chip, etc.

[0027] The detecting of lncRNA-92467 includes detecting the expression level of lncRNA-92467 in the pulmonary artery or cells thereof, preferably including extracting RNA in the pulmonary artery or cells thereof, reverse transcribing it into cDNA, and then performing PCR.

[0028] wherein the pulmonary artery or cells are derived from a subject.

[0029] The altitude sickness is an altitude sickness produced in a plateau environment.

[0030] The plateau environment is higher than 2000m, higher than 3000m, higher than 4000m, higher than 5000m or higher than 5500m. Preferably, the plateau environment is 2000-8000m above sea level, and more preferably, the plateau environment is 4000-6500m above sea level.

[0031] The plateau environment has low pressure and low oxygen conditions.

[0032] The altitude sickness may be acute altitude sickness or chronic altitude sickness.

[0033] The acute mountain sickness includes, but is not limited to, high altitude coma, high altitude cerebral edema, high altitude pulmonary edema, high altitude retinal hemorrhage, or a mixed disease in which abnormal brain and lung symptoms coexist.

[0034] The chronic altitude sickness includes, but is not limited to, a mixed disease in which altitude heart disease, altitude polycythemia, altitude hypertension, altitude hypotension, pulmonary hypertension, and altitude polycythemia coexist.

[0035] In some embodiments, the altitude sickness is pulmonary hypertension.

[0036] In some embodiments, the altitude sickness is altitude heart disease.

[0037] In some embodiments, the altitude sickness is pulmonary hypertension, ventricular hypertrophy (preferably right ventricular hypertrophy) or heart failure.

[0038] The heart failure mentioned herein refers to heart failure with reduced ejection fraction (HFrEF, LVEF<40%).

[0039] The early warning and / or diagnosis of altitude sickness includes detecting the expression level of lncRNA-92467 in the pulmonary artery or its cells, preferably including extracting RNA in the pulmonary artery or its cells, reverse transcribing it into cDNA, and then performing PCR.

[0040] Preferably, compared with a non-altitude disease patient or a threshold value, the expression level is significantly reduced, indicating that the patient is a patient with altitude disease or has a high probability of suffering from altitude disease.

[0041] In a second aspect, the invention provides the use of a lncRNA-92467 regulator in improving altitude tolerance or preparing a drug for treating and / or preventing altitude sickness.

[0042] The lncRNA-92467 regulator is an agent that upregulates lncRNA-92467 or an lncRNA-92467 inhibitor.

[0043] The reagent for upregulating lncRNA-92467 includes a reagent for upregulating the expression level of lncRNA-92467 or a reagent for improving the function of lncRNA-92467.

[0044] The lncRNA-92467 inhibitor is an agent that reduces the expression level of lncRNA-92467 or an agent that reduces the function of lncRNA-92467.

[0045] Preferably, the lncRNA-92467 regulator can be a small molecule compound, nucleic acid, protein, polypeptide, antibody, traditional Chinese medicine or its extract, interfering RNA or a reagent required for knocking out lncRNA-92467.

[0046] The interfering RNA includes but is not limited to one or a combination of two or more of siRNA, dsRNA, shRNA, aiRNA or miRNA, preferably siRNA.

[0047] The drug includes a lncRNA-92467 regulator and a pharmaceutically acceptable excipient.

[0048] Preferably, the pharmaceutically acceptable excipients include, but are not limited to, one or more of excipients, diluents, wetting agents, fillers, binders, lubricants, disintegrants, antioxidants, buffers, suspending agents, solubilizers, thickeners, stabilizers, flavoring agents and preservatives.

[0049] The drug can be administered by any suitable route, such as enteral administration (e.g. oral) or parenteral administration (e.g. intravenous, intramuscular, subcutaneous, intradermal, intraorgan, intranasal, intraocular, instillation, intracerebral, intrathecal, transdermal, intrarectal, etc.).

[0050] The drug can be in any suitable dosage form, such as a dosage form for gastrointestinal administration or a dosage form for parenteral administration, preferably including but not limited to tablets, pills, powders, granules, capsules, lozenges, syrups, liquids, emulsions, microemulsions, suspensions, injections, sprays, aerosols, powder sprays, lotions, ointments, plasters, pastes, patches, eye drops, nasal drops, sublingual tablets, suppositories, effervescent tablets, pills, gels, etc.

[0051] The various dosage forms of the drug can be prepared according to conventional production methods in the pharmaceutical field.

[0052] The drug may contain 0.01-99.5% by weight (specifically, 0.01%, 0.1%, 0.5%, 1%, 2%, 3%, 4%, 5%, 6%, 7%, 8%, 9%, 10%, 20%, 30%, 40%, 50%, 60%, 70%, 80%, 90%, 95%, 99%, 99.5%) of the lncRNA-92467 regulator.

[0053] In a third aspect, cells, tissues or organs that regulate the expression level of lncRNA-92467 are provided.

[0054] The regulation is either reduction or upregulation.

[0055] The cells are pulmonary artery endothelial cells.

[0056] The tissue is lung tissue.

[0057] The organ is the lung.

[0058] The expression level of lncRNA-92467 in the cells, tissues or organs is upregulated.

[0059] The expression level of lncRNA-92467 in the cell, tissue or organ is reduced, preferably knocked out or knocked down.

[0060] In a fourth aspect, a method for constructing cells, tissues or organs that regulate the expression level of lncRNA-92467 is provided.

[0061] The regulation is up-regulation or down-regulation.

[0062] The method for constructing cells, tissues or organs for upregulating the expression level of lncRNA-92467 comprises introducing a vector containing lncRNA-92467 into cells, tissues or organs.

[0063] The method for constructing cells, tissues or organs that reduce the expression level of lncRNA-92467 includes knocking out or knocking down lncRNA-92467 in cells, tissues or organs.

[0064] In a fifth aspect, a diagnostic kit is provided.

[0065] The diagnostic kit includes a reagent for detecting lncRNA-92467.

[0066] The diagnostic kit can diagnose or warn of altitude sickness.

[0067] In a sixth aspect, a method for warning and / or diagnosing altitude sickness is provided.

[0068] The method comprises detecting lncRNA-92467, preferably detecting the expression level of lncRNA-92467. It is further preferred to detect lncRNA-92467 in the pulmonary artery or its cells.

[0069] The detection method can adopt polymerase chain reaction (PCR), Southern blot hybridization, Northern blot hybridization, dot hybridization, fluorescence in situ hybridization (FISH), DNA microarray, high-throughput sequencing platform, chip detection, etc., especially PCR method, such as real-time fluorescence quantitative PCR method, constant temperature amplification technology (rolling circle nucleic acid amplification, loop-mediated isothermal amplification, chain displacement amplification, etc.).

[0070] In some embodiments, the method comprises extracting RNA from the pulmonary artery or cells thereof, reverse transcribing the RNA into cDNA, and then performing PCR.

[0071] The method comprises performing diagnosis using the diagnostic kit.

[0072] Preferably, compared with a non-altitude disease patient or a threshold value, the expression level is significantly reduced, indicating that the patient is a patient with altitude disease or has a high probability of suffering from altitude disease.

[0073] In a seventh aspect, a drug for treating and / or preventing altitude sickness is provided.

[0074] The drug includes a lncRNA-92467 regulator, such as an agent that upregulates lncRNA-92467 or a lncRNA-92467 inhibitor.

[0075] Preferably, a pharmaceutically acceptable excipient is also included.

[0076] The drug is for human use or veterinary use.

[0077] In an eighth aspect, a method for treating and / or preventing altitude sickness is provided.

[0078] The method comprises administering a lncRNA-92467 modulator or a cell or the drug that modulates the expression level of lncRNA-92467 to a subject in need thereof.

[0079] The method comprises introducing a lncRNA-92467 regulator or a cell that regulates the expression level of lncRNA-92467 or the drug into a subject.

[0080] The administration method can be enteral administration (e.g. oral) or parenteral administration (e.g. intravenous, intramuscular, subcutaneous, intradermal, intraorgan, intranasal, intraocular, instillation, intracerebral, intrathecal, transdermal, intrarectal, etc.) routes.

[0081] Ninthly, a method for altitude tolerance is provided.

[0082] The method comprises administering a lncRNA-92467 modulator or a cell or the drug that modulates the expression level of lncRNA-92467 to a subject in need thereof.

[0083] The method comprises introducing a lncRNA-92467 regulator or a cell that regulates the expression level of lncRNA-92467 or the drug into a subject.

[0084] The administration method can be enteral administration (e.g. oral) or parenteral administration (e.g. intravenous, intramuscular, subcutaneous, intradermal, intraorgan, intranasal, intraocular, instillation, intracerebral, intrathecal, transdermal, intrarectal, etc.) routes.

[0085] In a tenth aspect, a disease animal model is provided, wherein the disease animal model comprises introducing a lncRNA-92467 inhibitor into a non-human animal (eg, a cell, a tissue, or an organ).

[0086] The disease animal model is an animal model of altitude sickness, preferably an animal model of pulmonary hypertension.

[0087] The term "diagnosis" as used herein refers to finding out whether a patient has had a disease or condition in the past or at the time of diagnosis, or to finding out the progression of a disease.

[0088] The "early warning" mentioned in the present invention refers to finding out whether the patient is likely to get sick or have symptoms in the future, or finding out the possible future progression of the disease.

[0089] As used herein, "treating" means slowing, interrupting, preventing, controlling, stopping, alleviating, or reversing the progression or severity of a sign, symptom, disorder, condition, or disease after the disease has begun to develop, but does not necessarily involve the complete elimination of all disease-related signs, symptoms, conditions, or disorders.

[0090] The term "prevention" as used herein refers to a method implemented to prevent or delay the occurrence of a disease, disorder or symptom in an organism.

[0091] The "effective amount" described in the present invention refers to the amount or dosage of the drug of the present invention that provides the desired treatment or prevention after being administered to a subject in a single or multiple doses.

[0092] The "subject" described in the present invention can be a human or non-human mammal, or a cell, tissue or organ of a human or non-human mammal, and the non-human mammal can be a wild animal, a zoo animal, an economic animal, a pet, an experimental animal, etc. Preferably, the non-human mammal includes but is not limited to pigs, cattle, sheep, horses, donkeys, foxes, raccoon dogs, minks, camels, dogs, cats, rabbits, mice (such as rats, mice, guinea pigs, hamsters, gerbils, chinchillas, squirrels) or monkeys, etc. BRIEF DESCRIPTION OF THE DRAWINGS

[0093] The embodiments of the present invention are described in detail below with reference to the accompanying drawings, wherein:

[0094] Figure 1 :ceRNA network.

[0095] Figure 2 : HPAECs showed significant proliferation after being cultured under low-pressure and low-oxygen conditions. Nor was the normobaric and normoxic group, Hyp was the low-pressure and low-oxygen group, and β-actin was the internal reference.

[0096] Figure 3 : Relative expression level of lncRNA-92467 after HPAEC culture under hypobaric and hypoxic conditions, where Nor is the normobaric and normoxic group, Hyp is the hypobaric and hypoxic group, *** represents p < 0.001.

[0097] Figure 4 : Model construction results, where A is the result of red blood cell (RBCs) counting, B is the result of hemoglobin (HGB) concentration, C is the result of hematocrit (HCT), D is the result of mean pulmonary artery pressure (mPAP), E is the result of right ventricular systolic pressure (RVSP), F is the result of right ventricular hypertrophy index (RVHI), ns means the difference is not significant, * means p<0.05, ** means p<0.01, *** means p<0.001.

[0098] Figure 5 : The results of immunofluorescence and immunohistochemical staining were used to evaluate the thickness of arterial media, the degree of vascular muscularization and the cell proliferation rate.

[0099] Figure 6 : Relative expression level of lncRNA-92467 in rat pulmonary artery under a simulated plateau environment of 5500 meters above sea level, where Nor is the normobaric and normoxic group, Hyp is the hypobaric and hypoxic group, and ** represents p<0.01.

[0100] Figure 7: Effect of knockdown of lncRNA-92467 on HPAECs cell proliferation, where Nor is the normobaric and normoxic group, Hyp is the hypobaric and hypoxic group, Hyp+NC is the negative control hypobaric and hypoxic group, Hyp+si92467 is the hypobaric and hypoxic group after knockdown of lncRNA-92467, and β-actin is the internal reference.

[0101] Figure 8 : Effect of knockdown of lncRNA-92467 on HPAECs apoptosis, where Nor is the normobaric and normoxic group, Hyp is the hypobaric and hypoxic group, Hyp+NC is the negative control hypobaric and hypoxic group, Hyp+si92467 is the hypobaric and hypoxic group after knockdown of lncRNA-92467, and β-actin is the internal reference.

[0102] Fig. 9 : Effect of knocking down lncRNA-92467 on HPAECs cell migration, Nor is the normobaric and normoxic group, Hyp is the hypobaric and hypoxic group, Hyp+NC is the negative control hypobaric and hypoxic group, Hyp+si92467 is the hypobaric and hypoxic group after knocking down lncRNA-92467, the upper row of figures shows the cell migration results at 0 hours, and the lower row of figures shows the cell migration results at 24 hours. DETAILED DESCRIPTION

[0103] The following will be combined with the drawings in the embodiments of the present invention to clearly and completely describe the technical solutions in the embodiments of the present invention. Obviously, the described embodiments are only some embodiments of the present invention, not all. Based on the embodiments of the present invention, all other embodiments obtained by ordinary technicians in this field without making creative work are within the scope of protection of the present invention.

[0104] Animals used in the examples: SD rats, about 250 g, male, clean grade, purchased from Beijing Weitonglihua Experimental Animal Co., Ltd., license number: SCXK (Beijing) 2016-0006.

[0105] Cells used in the examples: pulmonary artery endothelial cells (HPAECs), human cells, purchased from Shanghai Guandao Biotechnology Co., Ltd.

[0106] In the embodiment, the hypobaric oxygen chamber is used for feeding: feeding with complete nutritional feed, the room temperature is 22-25° C., and the humidity is 30%-50%.

[0107] The instruments used in the examples are: small animal hypobaric oxygen chamber (Binglun Hyperbaric Oxygen Chamber Co., Ltd.); small animal ventilator (kent scientific); multifunctional physiological instrument (Millar); small animal ultrasound (Visual Sonics Inc); personalized cell precision control system (Avatar); Q-PCR instrument (Bio-Rad).

[0108] In the embodiment, the pulmonary artery of the animal was extracted: 3% sodium pentobarbital, 0.2 ml / 100 g, was intraperitoneally injected into the rat to anesthetize it, and ultrasonic detection was performed. The rat was then fixed on the operating table in a supine position, the trachea was cut, the ventilator was connected, the chest was opened, the heart was exposed, the catheter was inserted into the right ventricle, and the right heart catheter was tested. The rat was killed, the heart was taken out, the atrial tissue and attached fat were removed, the left and right ventricles were separated, the lung tissue was taken, the water was absorbed by filter paper, and the weights were respectively weighed, and the pulmonary artery was separated.

[0109] Statistical methods: All data were expressed as x±s, and the inter-group comparisons were performed using one-way analysis of variance. p<0.05 was considered statistically significant, and SPSS19.0 software package was used for statistical analysis.

[0110] Example 1: Marker screening

[0111] Through KEGG enrichment analysis, we selected mRNAs differentially expressed in HIF-1, PI3K-Akt, Wnt and other signaling pathways, performed lncRNA-mRNA co-expression analysis based on the data, and predicted miRNA-lncRNA and miRNA-mRNA relationship pairs based on the database to construct the ceRNA network (see Figure 1 ), screen differential lncRNAs. The specific steps are as follows:

[0112] The ceRNA network was predicted and constructed by Cytoscape version 3.6.0 software, and 19 lncRNA nodes were screened. Identification of hub genes in the ceRNA network associated with pulmonary hypertension. The topological characteristics of the ceRNA network, including PCC values ​​and degree distribution, were analyzed using Cytoscape software. Higher PCC values ​​in nodes indicate increased importance of these nodes in the regulatory network. From the hub network, four upregulated DElncRNAs (MSTRG.14380.1, ENSRNOT00000092424, ENSRNOT00000088836, MSTRG.19173.1) and two downregulated DElncRNAs (MSTRG.50957.1, ENSRNOT00000092467) were found with higher degree distribution and PCC values, indicating that they may be key regulators associated with pulmonary hypertension in the ceRNA network.

[0113] Further analysis of tyrosine phosphatase receptor M expression revealed that it was positively correlated with the hub lncRNA ENSRNOT00000092467 and played a role through cell adhesion molecules and adhesion junction signaling pathways. In addition, GO pathway enrichment indicated that tyrosine phosphatase receptor M was involved in a variety of biological processes, including endothelial cell proliferation, positive regulation of apoptotic cell clearance, cell differentiation, and cell development processes.

[0114] Example 2: Cell experiment on lncRNA-92467 expression level

[0115] 1. Experimental design

[0116] Pulmonary artery endothelial cells (HPAEC) were cultured in DMEM medium containing 10% fetal bovine serum and randomly divided into two groups. One group was placed in an individualized cell precision control system to adjust the oxygen concentration to 1% and the oxygen partial pressure to 0.6PSI to simulate the low-pressure and low-oxygen environment of the plateau. Different groups of cells were cultured in a normal-pressure and normoxic CO2 cell culture incubator for 24 hours.

[0117] 2. Detection method

[0118] Aspirate the cell culture medium, wash the cells twice with ice-cold PBS, extract RNA according to the requirements of the RNA extraction kit, and measure the concentration. According to the requirements of the RNA reverse transcription kit, reverse transcribe the RNA into cDNA and store it at -80°C. After diluting the cDNA 10 times, establish a 20ul system according to the requirements of the Q-PCR kit, set up the standard reaction mode and dissolution curve, and run the program.

[0119] 3. Results

[0120] After HPAEC cells were treated with hypotension and hypoxia for 24 hours, PCNA protein expression increased significantly, indicating that HPAEC cells proliferated significantly, that is, pulmonary hypertension occurred. Figure 2 shown.

[0121] The differential expression results are as follows Figure 3 As shown, after HPAECs cells were cultured in a low-pressure and low-oxygen environment, the expression of lncRNA-92467 in HPAECs cells was downregulated, and the difference was significant.

[0122] Example 3: Animal experiment on lncRNA-92467 expression level

[0123] 1. Experimental design

[0124] Twenty rats were randomly divided into two groups, with 10 rats in each group. One group was placed in an experimental cabin, and the pressure in the cabin was adjusted to 380 mmHg to simulate the plateau environment of 5,500 meters above sea level (low-pressure and low-oxygen group). At the same time, the environment in which the rats were located was maintained at a 12:12-hour day and night alternation. Different groups of rats were placed in the same room under normal pressure and normoxic environment and raised for 28 days.

[0125] The results are as follows Figure 4-5 As shown, rats in the low-pressure and low-oxygen group developed symptoms of pulmonary hypertension, including increased hemoglobin, increased pulmonary circulation-right heart pressure, right ventricular hypertrophy, and arteriolar vascular remodeling. The details are as follows:

[0126] The red blood cell (RBCs) count and hemoglobin (HGB) concentration were tested by routine blood tests. The RBCs count, HGB concentration and HCT value of rats increased with the extension of low pressure and hypoxia time. The RBCs count began to increase significantly on the 14th day, and the HGB and HCT values ​​increased significantly on the third day. Figure 4 AC.

[0127] RVSP and mPAP were measured by hemodynamics. Figure 4 As shown in DE, the RVSP and mPAP of rats raised in a low-pressure and low-oxygen environment increased over time, and the mPAP on the 28th day was 32.1976±1.2643 mmHg (greater than 20 mHg), indicating that the pulmonary hypertension model was successfully established.

[0128] Furthermore, the degree of right ventricular hypertrophy was evaluated by measuring the right ventricular hypertrophy index (RVHI) by cardiac weighing. Figure 4 F, The RVHI of rats increased with the extension of hypobaric and hypoxic time, and significant differences began to appear on the 7th day.

[0129] In addition, immunofluorescence and immunohistochemical staining were used to evaluate the arterial media thickness (MWT), vascular muscularization, and cell proliferation rate. Compared with day 0, the MWT, vascular muscularization, and cell proliferation rate of rats fed in a hypobaric and hypoxic environment for 21 days were significantly increased (P<0.001). Figure 5 shown.

[0130] 2. Expression level

[0131] Detection Methods

[0132] Aspirate the animal tissue blocks, wash the cells twice with ice-cold PBS, extract RNA according to the requirements of the RNA extraction kit, and measure the concentration. According to the requirements of the RNA reverse transcription kit, reverse transcribe the RNA into cDNA and store it at -80°C. After diluting the cDNA 10 times, establish a 20ul system according to the requirements of the Q-PCR kit, set up the standard reaction mode and dissolution curve, and run the program.

[0133] The results are as follows Figure 6 As shown, compared with rats raised in a normobaric and normoxic environment, the expression of lncRNA-92467 in the pulmonary artery of rats in the hypobaric and hypoxic group simulating a plateau environment of 5500 meters above sea level was downregulated, and the difference was significant.

[0134] Example 4: Effects of knocking down lncRNA-92467 on HPAECs

[0135] 1. Experimental design

[0136] 1) Design siRNA to specifically knock down the expression level of long-chain lncRNA-92467;

[0137] 2) Pulmonary artery endothelial cells (HPAECs) were cultured in DMEM medium containing 10% fetal bovine serum and randomly divided into 4 groups. The first group (Nor) was cultured in a normobaric and normoxic CO2 cell culture incubator as a control; the second group (Hyp) was placed in an individualized cell precision control system, and the oxygen concentration was adjusted to 1% and the oxygen partial pressure was 0.6PSI to simulate the low-pressure and low-oxygen environment of the plateau, and the cells were cultured for 48 hours; the third group (Hyp+NC) was transfected with irrelevant sequences into HPAECs cells, and the cells were placed in an individualized cell precision control system, and the oxygen concentration was adjusted to 1% and the oxygen partial pressure was 0.6PSI to simulate the low-pressure and low-oxygen environment of the plateau, and the cells were cultured for 48 hours; the fourth group (Hyp+siRNA) was transfected with the designed specific siRNA into HPAECs cells, and the cells were placed in an individualized cell precision control system, and the oxygen concentration was adjusted to 1% and the oxygen partial pressure was 0.6PSI to simulate the low-pressure and low-oxygen environment of the plateau, and the cells were cultured for 48 hours.

[0138] 2. Detection method

[0139] 1) Proliferation detection of the expression level of cell cycle-related protein PCNA;

[0140] 2) Migration detection method

[0141] After each group of HPAECs cells were cultured, the tip of the pipette was used to draw lines perpendicular to the cell plate, the cells were washed 3 times with PBS, and cell culture medium was added. Samples were taken at 0 and 24 h, and photos were taken. After opening the pictures with Image J software, the cell migration area was calculated.

[0142] 3) Apoptosis detection: the expression levels of Caspase3 and Caspase9.

[0143] 3. Results

[0144] The results are as follows Figure 7-9 As shown, Figure 7 This indicates that knocking down lncRNA-92467 can significantly promote HPAECs cell proliferation. Figure 8 The results showed that knockdown of lncRNA-92467 could significantly inhibit the apoptosis of HPAECs. Fig. 9 It showed that knocking down lncRNA-92467 could significantly promote HPAECs cell migration.

[0145] The preferred embodiments of the present invention are described in detail above. However, the present invention is not limited to the specific details in the above embodiments. Within the technical concept of the present invention, a variety of simple modifications can be made to the technical solution of the present invention, and these simple modifications all belong to the protection scope of the present invention.

[0146] It should also be noted that the various specific technical features described in the above specific embodiments can be combined in any suitable manner without contradiction. In order to avoid unnecessary repetition, the present invention will not further describe various possible combinations.

Claims

1. Application of lncRNA-92467 as a marker in the preparation of products for early warning and / or diagnosis of altitude sickness.

2. The use according to claim 1, characterized in that: The ID of the lncRNA-92467 is ENSRNOT00000092467.

3. The use according to claim 1, characterized in that: The product includes reagents for detecting the expression level of lncRNA-92467; Preferably, the reagents include reagents required for reverse transcription, reagents required for PCR, reagents required for sequencing, reagents required for mass spectrometry and / or buffer.

4. The use according to claim 1 or 3, characterized in that: lncRNA-92467 is lncRNA-92467 in pulmonary artery or its cells; Preferably, the cells are pulmonary artery endothelial cells.

5. The use according to claim 1, characterized in that: The altitude sickness is an altitude sickness produced in a plateau environment, and the plateau environment is an altitude higher than 2000m. Preferably, the plateau environment is an altitude of 2000-8000m, and more preferably, the plateau environment is an altitude of 4000-6500m, with low pressure and low oxygen conditions.

6. The use according to claim 1, characterized in that: The altitude sickness is acute altitude sickness or chronic altitude sickness; Preferably, the acute mountain sickness includes, but is not limited to, high altitude coma, high altitude cerebral edema, high altitude pulmonary edema, high altitude retinal hemorrhage, or a mixed disease with the coexistence of abnormal brain and lung symptoms; Preferably, the chronic altitude sickness includes, but is not limited to, a mixed disease in which altitude heart disease, altitude polycythemia, altitude hypertension, altitude hypotension, pulmonary hypertension, and altitude polycythemia coexist.

7. The use according to claim 1, characterized in that: The altitude sickness is pulmonary hypertension, ventricular hypertrophy or heart failure.

8. The use according to any one of claims 1 to 7, characterized in that: The early warning and / or diagnosis of altitude sickness comprises detecting the expression level of lncRNA-92467 in the pulmonary artery or its cells; Preferably, compared with a non-altitude disease patient or a threshold value, the expression level is significantly reduced, indicating that the patient is a patient with altitude disease or has a high probability of suffering from altitude disease.

9. The use according to claim 8, characterized in that: Detecting the expression level of lncRNA-92467 in the pulmonary artery or its cells includes extracting RNA in the pulmonary artery or its cells, reverse transcribing it into cDNA, and then performing PCR.

10. Use of lncRNA-92467 regulators in the preparation of drugs for treating and / or preventing altitude sickness.

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