CN109082465A - The molecular marker and application thereof of Olanzapine induction carbohydrate metabolism disturbance related disease - Google Patents
The molecular marker and application thereof of Olanzapine induction carbohydrate metabolism disturbance related disease Download PDFInfo
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Abstract
The invention discloses a kind of molecular markers of Olanzapine induction carbohydrate metabolism disturbance related disease, are the expression product of glucose transporter gene and/or glucose transporter gene.Glucose transporter gene and its expression product are related to the carbohydrate metabolism disturbance conspicuousness that Olanzapine induces, the action target spot of carbohydrate metabolism disturbance related disease can be induced as Olanzapine, be conducive to induce carbohydrate metabolism disturbance to carry out early diagnosis and immunotherapy targeted autoantibody Olanzapine, reduce the onset risk of Olanzapine drug induccd carbohydrate metabolism disturbance related disease.The invention discloses the screening techniques of the molecular marker of Olanzapine induction carbohydrate metabolism disturbance, take into account glucose uptake and utilization ways, improve the specific aim of molecular marker screening.The animal model of the Olanzapine induction carbohydrate metabolism disturbance constructed in the screening process of molecular marker, every glycometabolism index generate stability difference, provide reliable animal model for the screening of subsequent molecular marker and disease research.
Description
Technical field
The present invention relates to biomedicine fields, and in particular to the molecular marker of Olanzapine induction carbohydrate metabolism disturbance related disease
The screening technique of object and application thereof and the molecular marker of Olanzapine induction carbohydrate metabolism disturbance related disease.
Background technique
Schizophrenia (schizophrenia, SZ) is the high chronic delay of a kind of high recurrence rate, the disability rate of psychiatric department
Property disease, is one of the main reason for person between twenty and fifty are disabled.Clinical symptoms can be divided into cacesthesia (photis and phonism), vain hope,
The positive symptoms such as the chaotic mode of thinking, words and deeds are out of control, the feminine genders such as motivation lacks, floaty euphoria is reduced, social decline, aboulia
Symptom and attention are difficult to the cognitive defect concentrated, implementation capacity declines and working memory declines.Epidemiological survey display essence
The lifetime prevalence of refreshing Split disease worldwide is 3.8 ‰ -8.4 ‰ or so, is brought to patient, family members and society huge
Burden.
The new era of schizophrenia drug treatment has been started in nineteen fifty-two, the appearance of chlorpromazine, and antipsychotics starts
As schizoid main means are treated, occur in succession with perphenazine, Sulpiride, haloperidol etc. for representative later
First generation antipsychotics and be the second generation antipsychotic drug of representative with Clozapine, Olanzapine, Quetiapine, Risperidone etc.
Object.Compared with first generation antipsychotics, the action spectrum of second generation antipsychotics is wider, not only to schizoid
Positive symptom is effective, and is better than first to the remission effect of schizoid negative symptoms and its affective symptom to be occurred together
For drug；Current research prompt, second generation antipsychotic drug has slight improvement cognition to act on, and first generation antipsychotic drug
No this effect；In addition, extrapyramidal system adverse reaction (the extrapyramidal side- of second generation antipsychotics
Effects) less, drug safety is higher.Therefore, the recipe quantity of second generation antipsychotic drug is very big.
The clinical application of second generation antipsychotics is extensive, it was verified however that second generation antipsychotics easily causes
Weight gain, pathoglycemia etc. are metabolized relevant adverse reaction, induce schizophreniac's cardiovascular and cerebrovascular disease
The generation of (cardiovascular disease, CVD) seriously reduces the quality of life and treatment satisfaction of patient, influences
The compliance of patient and lapsing to for disease, easily lead to schizoid recurrence.Olanzapine is second generation antipsychotic
Recipe quantity is biggish in drug represents drug, the study found that taking the patient of Olanzapine, the incidence of cardiovascular and cerebrovascular disease for a long time
It is 2-3 times of general population.The risk that Olanzapine induces carbohydrate metabolism disturbance has been confirmed, and clearly marks in package insert
Show " receive the patient of olanzapine in treatment, glycemic control situation should be inspected periodically, measures fasting blood-glucose when starting treatment, and
Results of regular determination during treatment, and should be noted that the symptom for checking hyperglycemia, including polydipsia, diuresis, more foods, weakness ".
Therefore, improve carbohydrate metabolism disturbance caused by Olanzapine, the clinical therapeutic efficacy for improving Olanzapine has important
Meaning.But since the mechanism that Olanzapine causes carbohydrate metabolism disturbance is still not clear, limit disorderly to the glycometabolism of Olanzapine induction
Unrest and its improvement of related disease and treatment.
Summary of the invention
Therefore, the technical problem to be solved in the present invention is that overcoming the glycometabolism in the prior art for lacking Olanzapine induction
The therapy target information of disorder related disease leads to not carry out lacking for immunotherapy targeted autoantibody to the carbohydrate metabolism disturbance that Olanzapine induces
It falls into.
For this purpose, the invention provides the following technical scheme:
The present invention provides a kind of molecular marker of Olanzapine induction carbohydrate metabolism disturbance related disease, the molecular markers
Object is the expression product of glucose transporter gene and/or glucose transporter gene.
Optionally, above-mentioned molecular marker, the molecular marker are mRNA and/or the Portugal of glucose transporter gene
Grape sugar transporters albumen.
Optionally, above-mentioned molecular marker, the glucose transporter are II class glucose transporter and/or IV class Portugal
Grape sugar transporters.
Optionally, above-mentioned molecular marker, the carbohydrate metabolism disturbance related disease are selected from weight gain, obesity, high blood
Sugar, diabetes or cardiovascular disease.
The present invention provides above-mentioned molecular markers in preparation treatment Olanzapine induction carbohydrate metabolism disturbance related disease
Purposes in drug.
The present invention provides above-mentioned molecular markers early stage preparing Olanzapine induction carbohydrate metabolism disturbance related disease
Purposes in diagnostic reagent.
The present invention provides a kind of screening technique of the molecular marker of Olanzapine induction carbohydrate metabolism disturbance related disease, packets
Include following steps:
(1) animal model of building Olanzapine induction carbohydrate metabolism disturbance；
(2) the glucose uptake approach of the animal model and the index of correlation of glucose utilization approach are detected；
(3) screening has the different expression index of statistical significance, and the as described Olanzapine induces carbohydrate metabolism disturbance phase
The molecular marker of related disorders.
Optionally, above-mentioned screening technique, the animal model that the step constructs Olanzapine induction carbohydrate metabolism disturbance include:
The male SD rat for choosing weight 180-220g, is randomly divided into medicine group and control group；
After a week, medicine group rat is gavaged adaptive feeding with Olanzapine 2.1mg/kg weight, and blank group rat is with equivalent
Distilled water gavages, each group rat ad lib normal diet, free water during gavaging；
It persistently gavages 56-63 days, the glycometabolism index of the medicine group rat and the animal groups rat, which generates, has system
Meter learns the significant difference of meaning, obtains the animal model of Olanzapine induction carbohydrate metabolism disturbance.
Optionally, above-mentioned screening technique, the time persistently gavaged are 63 days.
Optionally, above-mentioned screening technique, the glycometabolism index include food ration, weight, fasting blood-glucose, empty stomach pancreas islet
Element, insulin sensitivity index and insulin resistance stable state evaluation model index.
Technical solution of the present invention has the advantages that
1. the present invention provides the molecular marker of Olanzapine induction carbohydrate metabolism disturbance related disease, the molecular marker
For glucose transporter gene and/or the expression product of glucose transporter gene.
Olanzapine may be by influencing neurotransmitters and its receptors such as serotonin, dopamine, norepinephrine, histamine
Function and interfere and normally ingest and satiety center, whet the appetite, along with the sedation of antipsychotics itself, use
Body activities amount is reduced after medicine, and energy consumption reduces, and eventually leads to weight gain.Meanwhile the carbohydrate metabolism disturbance of Olanzapine induction
Audient's multifactor impact, such as: it insulin secretion and response, glucose uptake and utilizes.Currently, lacking Olanzapine induction sugar
The action target spot information of metabolic disorder related disease limits the diagnosis to Olanzapine induction carbohydrate metabolism disturbance, treatment.
The present invention pass through the study found that Olanzapine induce carbohydrate metabolism disturbance medicine group rat in, glucose transporter
Expression quantity conspicuousness reduction in medicine group rat of (glucose transporter, GLUT) gene, albumen, and glycogen closes
Significant change does not occur for the content of enzyme, illustrates that Olanzapine induction carbohydrate metabolism disturbance mainly passes through Olanzapine to glucose uptake
It influences to determine, and Olanzapine is lower to the influence degree of glucose utilization approach, glucose transporter gene and its expression product
(mRNA, albumen) is related to the carbohydrate metabolism disturbance conspicuousness that Olanzapine induces.
The expression product of glucose transporter gene and/or glucose transporter gene can induce sugared generation as Olanzapine
It thanks to the molecular marker of disorder related disease, induces the diagnosing and treating of carbohydrate metabolism disturbance related disease to provide effectively for Olanzapine
Action target spot, in order to Olanzapine induction carbohydrate metabolism disturbance related disease carry out immunotherapy targeted autoantibody, advantageously reduce nitrogen difficult to understand
The onset risk of flat drug induccd carbohydrate metabolism disturbance related disease, improves the life quality of schizophreniac, improves anti-spirit
The compliance of disease treatment and patient lapse to, and reduce palindromia.
2, the molecular marker of the carbohydrate metabolism disturbance related disease of Olanzapine induction provided by the invention, glucose transporter
For II class glucose transporter (GLUT2 albumen, GLUT2mRNA) and/or IV class glucose transporter (GLUT4 albumen,
GLUT4mRNA).GLUT2 albumen is distributed mainly on liver organization, and GLUT4 albumen is mainly in the skeletal muscle of insulin sensitivity and rouge
It is expressed in fat tissue.In the medicine group rat of Olanzapine induction carbohydrate metabolism disturbance, GLUT2 albumen in liver organization,
The expression of GLUT4 albumen, GLUT4mRNA in GLUT2mRNA, and fat and musculature reduces, and illustrates nitrogen difficult to understand
The carbohydrate metabolism disturbance of flat induction is significant related to the expression of GLUT4 and GLUT2 gene, using above-mentioned molecular marker as nitrogen difficult to understand
The therapy target of flat induction carbohydrate metabolism disturbance can be realized and control Olanzapine induction carbohydrate metabolism disturbance early diagnosis and specific aim
It treats, improves the life quality of schizophreniac.
3, molecular marker provided by the invention is in the early diagnosis for preparing Olanzapine induction carbohydrate metabolism disturbance related disease
Purposes in reagent.Since the abnormal carbohydrate metabolism of Olanzapine induction generally requires certain reaction time, the present invention is with glucose
Transporter gene and its expression product are conducive to as target spot by detection glucose transporter gene, the expression of albumen
The early diagnosis to abnormal carbohydrate metabolism is realized before carbohydrate metabolism disturbance related disease occurs, in time to the glycometabolism of Olanzapine induction
Disorder carries out specific aim medication and treatment, prevents the generation of carbohydrate metabolism disturbance related disease.
4, the screening technique of the molecular marker of Olanzapine induction carbohydrate metabolism disturbance related disease provided by the invention, simultaneously
The index of correlation of glucose uptake approach and glucose utilization approach is detected, the uptake and utilization of glucose is taken into account, mentions
The high specific aim of Olanzapine induction carbohydrate metabolism disturbance molecular marker screening, eliminates in glycometabolism approach and induces with Olanzapine
The unrelated molecular indexes of carbohydrate metabolism disturbance are conducive to improve to Olanzapine induction carbohydrate metabolism disturbance related disease diagnosing and treating
Specific aim.
5, the method for the animal model of building Olanzapine induction carbohydrate metabolism disturbance provided by the invention, to medicine group rat and
Control rats persistently gavage, wherein after starting to gavage Olanzapine, it is poor that glycometabolism index of correlation starts to generate medicine group rat
It is different, but during gavaging, the difference of glycometabolism index of correlation generates fluctuation, influences animal model for screening Olanzapine induction sugar
The molecular marker of metabolic disorder and the accuracy of other correlative studys.In construction method provided by the invention, in lasting filling
After taking 56-63 days, the indices such as food ration, weight, fasting blood-glucose, fasting insulin, insulin sensitivity index generate stabilization
Sex differernce induces the research of carbohydrate metabolism disturbance related disease to provide reliably for the screening of subsequent molecular marker and Olanzapine
Animal model.
Detailed description of the invention
It, below will be to specific in order to illustrate more clearly of the specific embodiment of the invention or technical solution in the prior art
Embodiment or attached drawing needed to be used in the description of the prior art be briefly described, it should be apparent that, it is described below
Attached drawing is some embodiments of the present invention, for those of ordinary skill in the art, before not making the creative labor
It puts, is also possible to obtain other drawings based on these drawings.
Fig. 1 is testing result figure of the Olanzapine to the expressing quantity of Glycogensynthase；
Fig. 2 is that Olanzapine quantifies figure to the testing result of the expressing quantity of Glycogensynthase；
Fig. 3 is testing result figure of the Olanzapine to the expressing quantity of GLUT2 and GLUT4；
Fig. 4 is that Olanzapine quantifies figure to the testing result of the expressing quantity of GLUT2 and GLUT4；
Fig. 5 is testing result figure of the Olanzapine to the mrna expression amount of GLUT2 and GLUT4；
Fig. 6 is that Olanzapine quantifies figure to the testing result of the mrna expression amount of GLUT2 and GLUT4；
Fig. 7 is the testing result figure that Olanzapine influences food ration；
Fig. 8 is the testing result figure that Olanzapine influences weight；
Fig. 9 is Olanzapine on the horizontal testing result figure influenced of fasting blood-glucose (FBG)；
Figure 10 is the testing result figure that Olanzapine influences insulin level.
Specific embodiment
The laboratory apparatus that following embodiments are related to is as follows:
Blood glucose meter (Roche, the U.S., model: brilliant type)；Assay balance (Shanghai Jingtian Electronic Instrument Co., Ltd., model:
FA2104A)；Centrifuge (the silent winged generation that of match, the U.S., model: MR23i)；Enzyme micro-plate reader (the limited public affairs of Shanghai China of section experimental system
Department, model: ST-360)；Ultraviolet-uisible spectrophotometer (Beckman, the U.S., model: DU730)；Microscope (Olympus,
Model: CX23)；Odyssey infrared imaging system imaging system (LI-COR, the U.S.)；Fluorescence quantitative PCR instrument (ABI, the U.S., type
Number: 7900).
The drug that following embodiments are related to and reagent are as follows:
Olanzapine Tablets (trade name: Ou Lanning, Jiangsu Haosen Pharmaceutical Co., Ltd, lot number: 171104, specification: 5mg/
Piece), grind powder after with distilled water dissolution Olanzapine suspension is made；(He Peng (Shanghai) is raw for rat insulin ELISA detection kit
Object Science and Technology Ltd., lot number 201712)；Phosphate buffer (PBS, pH7.4, Life technologies, the U.S., article No.
10010049)；GS antibody (AbCAM, Britain, article No. ab40810)；GLUT-2 antibody (SAB, the U.S., article No. 45050-1)；
GLUT-4 antibody (AbCAM, Britain, article No. ab188317)；GAPDH antibody (Abbkine, the U.S., article No. Abp57259)；Secondary antibody
IRDye 680CW (Licor, the U.S.)；Marker (the green skies Bioisystech Co., Ltd in Shanghai)；The examination of BCA determination of protein concentration
Agent box (Beijing Suo Laibao Science and Technology Ltd, article No. PC0020)；(health is that century biotechnology has to ultrapure RNA extracts kit
Limit company, article No. CW0581S)；(health is century Bioisystech Co., Ltd, goods to the first chain of SuperRT cDNA synthetic agent box
Number CW0741S)；Quantitative fluorescent PCR premixes system (health is century Bioisystech Co., Ltd, article No. CW2602M)；Primer synthesis
(Hua Da gene).
The male cleaning grade SD rat of the health that following embodiments use is purchased from the western limited public affairs of Poole-Bi Kai experimental animal in Shanghai
Department, quality certification number SCXK (Shanghai) 2013-0016.
Embodiment 1
The present embodiment provides the construction methods of the animal model of Olanzapine induction carbohydrate metabolism disturbance, comprising the following steps:
(1) healthy male cleaning grade SD rat 12,180~220g of weight, 6~8 week old, feeding environment: room temperature are chosen
20~26 DEG C, relative humidity 40%~70%, daily illumination 12h (07:00~19:00), ad lib normal diet are freely drunk
Water.12 rats are divided into medicine group and control group at random, and every group 6.
(2) after adaptive feeding 1 week, daily dose as defined in Consult drug specification converts, every afternoon 17:00,
Medicine group rat is gavaged with Olanzapine 2.1mg/kg weight, and blank group rat is gavaged with equal amount of distilled water.During experiment, each group is big
Mouse ad lib normal diet, free water,
(3) it persistently gavages 63 days, detects the glycometabolism index of medicine group rat and the animal groups rat, glycometabolism index
Refer to including food ration, weight, fasting blood-glucose, fasting insulin, insulin sensitivity index and insulin resistance stable state evaluation model
Number, glycometabolism index generate stable significant difference, obtain the animal mould for constructing successful Olanzapine induction carbohydrate metabolism disturbance
Type.
Embodiment 2
The present embodiment provides a kind of Olanzapine induction carbohydrate metabolism disturbance molecular marker screening technique, specifically include with
Lower step:
1, medicine group mouse and the control group mice of end will be administered in embodiment 1, using CO2Gas euthanasia method is put to death
Rat acquires liver, fat, musculature respectively, is weighed and placed in PBS, and 4 DEG C of preservations are used for subsequent Indexs measure.
2, the glucose uptake approach of the animal model and the index of correlation of glucose utilization approach are detected.Wherein, Portugal
The index of correlation of grape Sugar intake approach includes GLUT2 albumen, GLUT2mRNA, GLUT4 albumen and GLUT4mRNA, glucose utilization
The index of correlation of approach includes Glycogensynthase.
The measurement of 2.1 hepatic glycogen and muscle glycogen content
Liver organization to be measured is taken, is rinsed with physiological saline, filter paper blots, weighing, about 80mg or so, according to sample weight
(mg): lye volume (μ L)=1:3 is added in test tube, boiling water bath 20min together, and flowing water is cooling, and hepatic glycogen hydrolyzate is made,
Glycogenolysis liquid is further prepared into 1% hepatic glycogen detection liquid with distilled water, is measured according to kit specification and calculates liver
Glycogen content.Muscle glycogen content need to prepare 5% muscle glycogen detection liquid when measuring, remaining method is measured with hepatic glycogen content.
The detection of 2.2 Glycogensynthases (GS) activity and protein expression level
The activity of determined by ultraviolet spectrophotometry GS: it according to liver to be measured and musculature quality (about 0.1g or so), presses
Extracting liquid volume is added according to 1:10 ratio, is homogenized under condition of ice bath, 4 DEG C, 8 000 × g be centrifuged 10min, take supernatant, set on ice to
It surveys.Ultraviolet specrophotometer preheats 45min, adjusting wavelength 340nm, distilled water zeroing；It is added in quartz colorimetric utensil to test sample
Product and dielectric fluid mix；The absorbance A 2 for recording initial absorbance A1 and 1min at 340nm, according to the calculation formula of kit
Calculate the activity of GS.
The protein expression of Western Blot method measurement GS: taking liver to be measured and musculature, is control with GAPDH,
Transferring film after SDS-PAGE electrophoresis closes film with 1% bovine serum albumin(BSA) (BSA)；Add primary antibody and control antibodies, 4 DEG C incubate
It educates overnight；PBS is washed three times, adds secondary antibody IRDye680CW, 37 DEG C of incubation 1h；After PBS cleaning three times, clapped with fluoroscopic imaging systems
According to, read as a result, with the albumen relative expression quantity of destination protein and the ratio calculation GS of internal reference albumen.
The detection of 2.3 glucose transporters (GLUT) protein expression ability
The protein expression of Western Blot method measurement GLUT: taking test serum sample, is control, Western with GAPDH
Blot method measures the protein expression level of sugar GLUT, protein expression detection process of the specific method with GS.
The mRNA expression of PCR method measurement GLUT: test serum sample is taken, is control with β-actin, is mentioned using ultrapure RNA
It takes kit to extract total serum IgE, SuperRT cDNA the first chain synthetic agent box is used at cDNA, to use RNA reverse transcription after quantitative
The mRNA expression of fluorescence quantitative PCR instrument testing goal albumen.PCR amplification condition: 50 DEG C of 2min, 95 DEG C of 10min, 95 DEG C,
15s, 60 DEG C, 60s, 40 circulations.PCR primer sequence is as follows:
GLUT2-F, nucleotide sequence is as shown in SEQ ID NO.1；
GLUT2-R, nucleotide sequence is as shown in SEQ ID NO.2；
GLUT4-F, nucleotide sequence is as shown in SEQ ID NO.3；
GLUT4-R, nucleotide sequence is as shown in SEQ ID NO.4；
β-actin-F, nucleotide sequence is as shown in SEQ ID NO.5；
β-actin-R, nucleotide sequence is as shown in SEQ ID NO.6.
2.4 statistical method
Data are analyzed using SPSS18.0 statistical software, measurement data is indicated with mean ± standard deviation (s), multiple groups
Between compare using variance analysis, compare two-by-two between group and examined using LSD-t, it is statistically significant by difference of P0.05.
3, experimental result
The activity and expressing quantity testing result of 3.1 glycogens and enzyme:
Glycogen content is measured in rat liver and musculature after being induced for a long time Olanzapine respectively, as a result such as table
Shown in 1, the hepatic glycogen content of medicine group rat is reduced compared with control group, and difference is statistically significant (P < 0.05)；Muscle glycogen content
Also it reduces, but no significant difference (P > 0.05).
Table 1 gives influence of the Olanzapine to glycogen content for a long time
| Group | Hepatic glycogen (mg/g) | Muscle glycogen (mg/g) |
| Control group | 6.37±0.45 | 1.35±0.05 |
| Medicine group | 5.95±0.17* | 1.25±0.06 |
Note: compared with the control group, * P < 0.05
Fig. 1 and Fig. 2 shows testing result figure of the Olanzapine to the expressing quantity of Glycogensynthase, Fig. 1 western-
Blot testing result, Fig. 2 be Fig. 1 quantized result figure, by Fig. 1 and Fig. 2 it is found that Olanzapine induce for a long time after rat liver with
There is no large change (P > 0.05) for the expressing quantity of Glycogensynthase in musculature.
The active testing result of Glycogensynthase such as table 2 in rat liver and musculature after being induced for a long time Olanzapine
Shown: Glycogen synthase activity increases compared with control group in medicine group rat liver, and difference is statistically significant (P < 0.05)；Muscle
Middle Glycogen synthase activity also increases, but no significant difference (P > 0.05).
Table 2 gives influence of the Olanzapine to Glycogen synthase activity for a long time
Note: compared with the control group, * P < 0.05
From the above results, the expression of Glycogensynthase and activity do not drop in body after Olanzapine induces for a long time
It is low, illustrate under being induced for a long time due to Olanzapine, the synthesis capability of glycogen is not affected, and Olanzapine is to glucose using not
Generation significantly affects.
The testing result of protein expression and the mRNA expression of 3.2 glucose transporters
Fig. 3 and Fig. 4 shows Olanzapine to the testing result figure of the expressing quantity of GLUT2 and GLUT4, and Fig. 3 is
Western-blot testing result, Fig. 4 are the quantized result figure of Fig. 3；Fig. 5 and Fig. 6 shows Olanzapine to GLUT2's and GLUT4
The testing result figure of mrna expression amount, Fig. 5 are agarose gel electrophoresis testing result, and Fig. 6 is the quantized result figure of Fig. 5.By scheming
3- Fig. 6 it is found that in rat liver tissue after being induced for a long time Olanzapine respectively in GLUT2, muscle and adipose tissue GLUT4 into
Row detection discovery, in observed tissue, the protein expression and mRNA of medicine group GLUT, which is expressed, to be declined compared with control group, difference
There is height statistical significance (P < 0.01).
The above results show after giving Olanzapine for a long time that the expressing quantity of Glycogensynthase does not have in liver and musculature
It changes, protein active enhancing, under showing that Olanzapine induces for a long time, Glycogen synthesis is not affected.In conjunction with the present invention
The research conclusion of middle medicine group glycogen content reduction can speculate and show that the ability of Olanzapine increase decomposition of glycogen is far longer than rush
Into the ability of Glycogen synthesis, thus cause blood glucose fluctuation.On the other hand, by detect give for a long time rat liver after Olanzapine,
The variation of glucose transporter expression is found in muscle, adipose tissue, the albumen and mRNA table of glucose transporter in each tissue
Decline up to amount, thus speculates, Olanzapine influences suction of the cell to glucose by inhibiting the normal ingestion of glucose to transport
It receives, increases the concentration of glucose in blood.
The studies above the result shows that, under the long-term induction of Olanzapine, body appetite and weight gain, glucose in tissue
Transporter expression, which is lowered, causes cellular uptake glucose ability limited, and decomposition of glycogen increase destroys sugared stable state in vivo, generates pancreas
Insulin resistance, final the concentration of glucose in blood are higher than normal value.Blood glucose rise is the potential risk of New-Onset Diabetes Mellitus, Olanzapine
Drug induccd hyperglycemia not only threatens the cardiovascular health of drug user, also therefore can reduce the treatment satisfaction of patient, influence
Compliance and disease lapse to, and easily lead to the recurrence of schizophrenia disease, increase the negative of personal, family and society
Load.
Key effect of the present invention by research discovery GLUT2 and GLUT4 in the abnormal carbohydrate metabolism that Olanzapine induces, Portugal
Grape sugar transporters (glucose transporter, GLUT) albumen is a kind of transmembrane transporter being present on cell membrane, is born
The glucose transport of intraor extracellular is blamed, hydrophily glucose cannot be alone across cell membrane, it is necessary to be by special transport protein
GLUT albumen is carried along into the cell, and the expression of GLUT reduces or activity change can influence histocyte to the intake energy of glucose
Power, the present invention by research confirm, Olanzapine induction carbohydrate metabolism disturbance in, Olanzapine by influence GLUT2 mRNA,
GLUT2 albumen, GLUT4 mRNA and GLUT4 albumen normal expression, cause glucose uptake difficult, make glucose in blood
Concentration increases, and leads to carbohydrate metabolism disturbance.Therefore, GLUT2 and GLUT4 can induce carbohydrate metabolism disturbance related disease as Olanzapine
Molecular marker.Molecular marker provided by the invention is provided for the abnormal carbohydrate metabolism that Olanzapine induces and new is effectively controlled
Target spot is treated and/or detected, effective information is provided for developing new drug object, implementation individualized clinical treatment scheme, is mental disease
Drug user brings benefit.
Experimental example 1
1, experiment purpose: the glycometabolism of the animal model of the Olanzapine induction carbohydrate metabolism disturbance constructed in detection embodiment 1
Index.
2, experimental method:
The measurement of 2.1 food rations, weight
The food ration of record each group rat daily, claims weekly a weight.
The measurement of 2.2 fasting blood-glucoses (FBG) level
It is horizontal that a fasting blood-glucose (FBG) is surveyed weekly, after fasting 12h, by 10 μ L of tail vein blood, according to Roche blood glucose
The operating instruction of instrument measures FBG value.
2.3 Olanzapines are to insulin level, insulin sensitivity index (ISI) and insulin resistance stable state evaluation model index
(HOMA-IR) influence measurement
1, the experimental material and experimental material provided with embodiment 1 obtains medicine group mouse and the control group of Olanzapine processing
Mouse.
2, the measurement of fasting insulin (FINS) level and insulin sensitivity index (ISI) and insulin resistance stable state are commented
The calculating of valence model index (HOMA-IR):
It is horizontal that a fasting insulin (FINS) is surveyed weekly: after fasting 12h, by 100 μ L of tail vein blood, being stood
30min, centrifugation take serum, and ELISA method measures FINS value.Detailed process includes:
(1) it balances: lath needed for being taken out from the aluminium foil bag after equilibrium at room temperature 20min；
(2) standard sample wells is arranged: standard sample wells respectively adds the 50 μ L of standard items of various concentration；
(3) it is loaded: in sample to be tested hole, first plus 40 μ L of sample diluting liquid, then again plus 10 μ L of sample to be tested, shaking, is mixed
It is even；
(4) enzyme: 100 μ of detection antibody that horseradish peroxidase (HRP) is marked is added in standard sample wells and the every hole of sample aperture
L seals reacting hole, 37 DEG C of incubation 60min with sealing plate film；
(5) it is washed with liquid: will be spare after 20 times of concentrated cleaning solutions, 20 times of distilled water dilutions；
(6) wash: carefully taking sealing plate film off, discard liquid, dry, cleaning solution is filled it up in every hole, standing 1min, after discard,
It is repeated 5 times, pats dry；
(7) develop the color: color developing agent A50 μ L is first added in every hole, adds color developing agent B50 μ L, and concussion mixes, 37 DEG C be protected from light it is aobvious
Color 15min；
(8) terminate: every hole adds 50 μ L of terminate liquid, terminates reaction；
(9) it measures: being returned to zero with blank well, 450nm wavelength sequentially measures the absorbance (OD value) in each hole, and measurement is adding termination
It is carried out in 15min after liquid.(11) calculate: using the concentration of standard items as abscissa, OD value is ordinate, calculates standard curve, will
The OD value of sample substitutes into standard curve, sample concentration is calculated, multiplied by extension rate, the as actual concentrations (FINS of sample
Value).
Then, ISI the and HOMA-IR value of body after giving Olanzapine for a long time: ISI=LN is calculated separately by following equation
[1/(FBG*FINS)]；HOMA-IR=FBG*FINS/22.5.
3, experimental result:
Fig. 7 shows the testing result that Olanzapine influences food ration, and in experiment first 35 days, animal is in drug and stomach-filling is suitable
Answer character state, the food ration of two groups of rats in this stage in variation irregularities；After 35 days, medicine group food ration is integrally big
In control group.
Fig. 8 shows the testing result that Olanzapine influences weight, observes the variation hair of medicine group and control rats weight
It is existing, before experiment in 14 days, the basic indifference of the body weight increase of two groups of rats；After 14 days, the weight of medicine group rat is generally big
In control group, but the no significant difference in the stage；To experiment, weight and the control group phase of medicine group rat
Than producing the difference (P < 0.05) of statistical significance.
Fig. 9 shows that Olanzapine on the horizontal testing result influenced of fasting blood-glucose (FBG), is tested first 7 days, the sky of two groups of rats
Abdomen blood glucose has transient raising；After 7 days, the fasting blood-glucose of two groups of rats is overall on a declining curve；When 35 days, two groups
Fasting blood glucose level tends to be close, basicly stable level before administration；By 35 days drug adaptable fed stages, drug
Group rat fasting blood-glucose is horizontal whole higher than control group；At the 56th day, two groups statistically significant (P < 0.05) compared to difference；
At the 63rd day, two groups have height statistical significance (P < 0.01) compared to difference.
Figure 10 shows the testing result that Olanzapine influences insulin level, tests first 14 days, the insulin of two groups of rats
Horizontal is in the trend that persistently increases, and 0-7 days elevated-levels are greater than 7-14 days；After 14 days, the insulin level of two groups of rats is equal
Decline, this stage insulin level is unstable, irregularities；By 35 days drug adaptable fed stages, control rats
Insulin level be basically stable on similar level, no large change, and the insulin level of medicine group rat is integrally presented
The trend of liter；At the 63rd day, for medicine group compared with control group insulin level, difference is statistically significant (P < 0.05).According to
Formula calculates separately the insulin sensitivity index and insulin resistance stable state evaluation model index of two groups of rats, as a result such as 3 institute of table
Show: the insulin sensitivity index (ISI) of experiment the 63rd day, medicine group rat declines compared with control group, the statistically significant (P of difference
< 0.05)；The insulin resistance stable state evaluation model index (HOMA-IR) of medicine group is greater than control group simultaneously, and difference has height
Statistical significance (P < 0.01).
Table 3 gives influence of the Olanzapine to ISI and HOMA-IR for a long time
| Group | ISI | HOMA-IR |
| Control group | -5.71±0.10 | 13.46±1.32 |
| Medicine group | -6.00±0.09* | 17.95±1.57** |
Note: compared with the control group, * P < 0.05, * * P < 0.01
From the above results, after persistently gavaging Olanzapine 63 days, the food ration of medicine group rat and control rats,
Weight, FBG, FINS, ISI and HOMA-IR generate the change of divergence of regularity, and difference has statistical significance, success structure
The animal model for having built Olanzapine induction carbohydrate metabolism disturbance, the molecular marker of carbohydrate metabolism disturbance related disease is induced for Olanzapine
The disease research of screening and its Olanzapine induction carbohydrate metabolism disturbance provides reliable animal model.
Obviously, the above embodiments are merely examples for clarifying the description, and does not limit the embodiments.It is right
For those of ordinary skill in the art, can also make on the basis of the above description it is other it is various forms of variation or
It changes.There is no necessity and possibility to exhaust all the enbodiments.And it is extended from this it is obvious variation or
It changes still within the protection scope of the invention.
Sequence table
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Claims (10)
1. a kind of molecular marker of Olanzapine induction carbohydrate metabolism disturbance related disease, which is characterized in that the molecular marker
For glucose transporter gene and/or the expression product of glucose transporter gene.
2. molecular marker according to claim 1, which is characterized in that the molecular marker is glucose transporter base
The mRNA and/or glucose transport body protein of cause.
3. molecular marker according to claim 1 or 2, which is characterized in that the glucose transporter is II class grape
Sugar transporters and/or IV class glucose transporter.
4. molecular marker according to claim 1-3, which is characterized in that the carbohydrate metabolism disturbance related disease
Selected from weight gain, obesity, hyperglycemia, diabetes or cardiovascular disease.
5. the described in any item molecular markers of claim 1-4 induce carbohydrate metabolism disturbance related disease in preparation treatment Olanzapine
Drug in purposes.
6. the described in any item molecular markers of claim 1-4 are in the morning for preparing Olanzapine induction carbohydrate metabolism disturbance related disease
Purposes in phase diagnostic reagent.
7. a kind of screening technique of the molecular marker of Olanzapine induction carbohydrate metabolism disturbance related disease, which is characterized in that including
Following steps:
(1) animal model of building Olanzapine induction carbohydrate metabolism disturbance；
(2) the glucose uptake approach of the animal model and the index of correlation of glucose utilization approach are detected；
(3) screening has the different expression index of statistical significance, and the as described Olanzapine induces carbohydrate metabolism disturbance correlation disease
The molecular marker of disease.
8. screening technique according to claim 7, which is characterized in that the step building Olanzapine induces carbohydrate metabolism disturbance
Animal model include:
The male SD rat for choosing weight 180-220g, is randomly divided into medicine group and control group；
After a week, medicine group rat is gavaged adaptive feeding with Olanzapine 2.1mg/kg weight, and blank group rat is distilled with equivalent
Water gavages, each group rat ad lib normal diet, free water during gavaging；
It persistently gavages 56-63 days, the glycometabolism index of the medicine group rat and the animal groups rat, which generates, has statistics
The significant difference of meaning obtains the animal model of Olanzapine induction carbohydrate metabolism disturbance.
9. screening technique according to claim 8, which is characterized in that the time persistently gavaged is 63 days.
10. screening technique according to claim 8, which is characterized in that the glycometabolism index include food ration, weight,
Fasting blood-glucose, fasting insulin, insulin sensitivity index and insulin resistance stable state evaluation model index.
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