CN114392266A

CN114392266A - Pharmaceutical composition containing PPAR gamma inhibitor and application thereof

Info

Publication number: CN114392266A
Application number: CN202210075688.4A
Authority: CN
Inventors: 潘颖; 李瑞满; 刘海智; 韩小雪; 邱迪
Original assignee: First Affiliated Hospital of Jinan University
Current assignee: First Affiliated Hospital of Jinan University
Priority date: 2022-01-22
Filing date: 2022-01-22
Publication date: 2022-04-26
Anticipated expiration: 2042-01-22
Also published as: CN114392266B

Abstract

The invention provides a pharmaceutical composition containing a PPAR gamma inhibitor and used for promoting neuron differentiation and treating neuron differentiation disorder and application thereof. The invention clearly discloses the function and mechanism of PPAR gamma in the process of neuron differentiation under high-sugar environment. The PPAR gamma has a regulation effect with autophagy activity and can mediate the autophagy process, so that the neuron differentiation is inhibited, and the normal development of a nervous system is influenced; after the interference of the PPAR gamma, the expression of the PPAR gamma in the cell can be obviously reduced, and the autophagy process of the cell is obviously inhibited, so that the differentiation of the neuron is promoted. Discloses a new mechanism of PPAR gamma for regulating and controlling neuron differentiation under a high-sugar environment, and provides practical experimental evidence and research direction for clinically intervening and treating relevant neurological diseases caused by gestational diabetes.

Description

Pharmaceutical composition containing PPAR gamma inhibitor and application thereof

Technical Field

The invention belongs to the field of biological medicines, relates to a pharmaceutical composition containing a PPAR gamma inhibitor and application thereof, and particularly relates to a pharmaceutical composition containing a PPAR gamma inhibitor and used for promoting neuron differentiation and treating neuron differentiation disorder and application thereof.

Background

Diabetes is a group of metabolic syndromes caused by the interaction of various factors such as heredity and environment, and is mainly characterized by chronic hyperglycemia. Pregnancy complicated with diabetes is one of the most common medical complications in pregnancy, and long-term hyperglycemia causes systemic microangiopathy such as fundus and kidney; large blood vessel diseases such as heart and brain, and chronic progressive changes such as neuropathy. With the development of socio-economic, the number of patients with pregnancy complicated with diabetes is increasing.

The influence of gestational diabetes mellitus on pregnant women and fetuses is closely related to the time of the appearance of maternal blood sugar rise and the blood sugar level during pregnancy. Complications such as fetal congenital malformation, abortion, early growth restriction and the like may be caused in the early stage of pregnancy. Common fetal abnormalities are: neural tube deformity, heart deformity, kidney deformity, digestive tract deformity, cleft lip and palate, etc. In the middle and late period of pregnancy, giant infants, delayed lung development, delayed development of central nervous system, intrauterine stillbirth and the like may appear. Because the normal development of the brain of a fetus is disturbed due to the disturbance of maternal glucose metabolism in the gestation period, the brain of a newborn falls to the same age after the development and maturity degree, and various behavioral and intellectual disorders of the newborn can be seriously caused.

A large number of studies indicate that a series of problems occur in the nerve development process after birth, and scholars think that the consequences are caused by brain dysplasia and brain injury caused by exposure of children to the environment with glucose metabolism disorder during pregnancy, and most of the consequences are as follows: exercise lag, abnormal muscle tone, speech and eye movement dysfunction, poor social accommodation ability, inattention, memory impairment, and the like. After the adult, obesity and impaired glucose tolerance appear, and further diseases such as diabetes, metabolic syndrome and the like which seriously affect life appear.

Hyperglycemia often causes severe complications, for example, plays an important role in the development of diabetic neuropathy, nephropathy, and retinopathy. Diabetic neuropathy is neuronal damage caused by prolonged exposure to high glucose. Recent studies have shown that by 2030, approximately 2-3 million people will be expected to have diabetic neuropathy. Diabetic neuropathy is a multifunctional disease, and its pathogenesis is abnormally complex due to the simultaneous participation of multiple signaling pathways.

Autophagy (Autophagy) is an integral pathway for lysosomal degradation, and is involved in the recovery of longevity proteins and cytoplasmic organelles. It is negatively regulated by the serine/threonine kinase mTOR and is a key process in cell growth and metabolism, which ultimately produces macromolecules to maintain cellular homeostasis. Up to now, more than 30 autophagy-related (Atg) genes have been identified, which are highly conserved in eukaryotes, e.g., the Atg protein family, LPC-I, LPC-II, Beclin-1, p62, etc. In inducing autophagy, the conversion of Beclin, LC3-I to LC3-II indicates the formation of autophagosomes and is therefore widely used as a marker for autophagosome formation. This pathway depends to a large extent on the activity of the lysosome, and any defect in lysosomal degradation of the autophagosome can lead to its accumulation, impair cell function, and possibly lead to cell death through protease activation.

Neuronal differentiation is critical for fetal brain development, while autophagy levels severely affect neuronal differentiation from stem cells to neurons. Embryonic stem cells are pluripotent stem cells present in early embryos and have the ability to differentiate into the predominant germ layer. The regulation and control of autophagy level are critical to stem cell differentiation, and electron microscope observation shows that damaged autophagy bodies are formed when unhealthy stem cells are differentiated, which indicates that autophagy pathway is blocked and the volume is reduced, so that protein degradation is realized. Homozygous mutations in key genes such as the autophagy gene Ambra1 also cause embryonic lethality in mice, and their functional defects in embryonic stem cells result in severe neural tube defects, all of which are associated with autophagy dysfunction and dysregulation.

The existing research shows that the high sugar can inhibit the differentiation of the neural stem cells, but the specific molecular mechanism is not clear, and the role of autophagy activity in the differentiation process of the neural stem cells needs to be further researched. Therefore, the research and the explanation of the development and the progress of the diabetic neuropathy and the key molecular approach of the pathogenesis-oriented drug treatment target point have very important significance, and have important theoretical guidance function for clinically preventing the neuropathy caused by the gestational diabetes.

Disclosure of Invention

The invention aims to solve the problems in the prior art, thereby carrying out intensive research on related functions and mechanisms of hyperglycemia, neuron differentiation, diabetic neuropathy and the like in diabetes, particularly gestational diabetes, disclosing a new mechanism for effectively regulating neuron differentiation disorder caused by hyperglycemia by regulating and controlling the activity of a target spot PPAR gamma, and providing practical experimental evidence and scientific basis for clinical intervention and treatment of related neurological diseases caused by gestational diabetes.

In order to solve the above technical problems, the present invention is achieved by the following technical solutions.

In a first aspect, the present invention provides a pharmaceutical composition for promoting neuronal differentiation comprising a PPAR γ inhibitor.

Preferably, the PPAR γ inhibitor is selected from shRNA (shpara) designed based on PPAR γ gene.

Preferably, the shPPAR γ is selected from one or more of shPPAR γ -1, shPPAR γ -2 and shPPAR γ -3; the sequences of the shPPAR gamma-1, the shPPAR gamma-2 and the shPPAR gamma-3 are respectively shown as SEQ ID NO: 1. SEQ ID NO: 2. SEQ ID NO: 3, respectively.

In a second aspect, the invention provides a pharmaceutical composition for the treatment of a neuronal differentiation disorder comprising a PPAR γ inhibitor.

Preferably, the neuronal differentiation disorder is a diabetic neuronal differentiation disorder.

Preferably, the diabetic neuron differentiation disorder is a gestational diabetic neuron differentiation disorder.

In a third aspect, the invention provides the use of a PPAR γ inhibitor in the manufacture of a medicament for inhibiting hyperglycemia-induced autophagy.

In a fourth aspect, the invention provides the use of a PPAR γ inhibitor in the manufacture of a medicament for the treatment of a neuronal differentiation disorder.

In a fifth aspect, the present invention provides the use of a PPAR γ inhibitor in the manufacture of a medicament for promoting neuronal differentiation.

Peroxisome proliferator-activated receptors (PPARs) are a family of type II nuclear receptors, comprising three isoforms (α, β/δ and γ). PPARs are ubiquitously expressed in tissues, presenting cell type and developmental stage specificity. Their transcriptional activity is regulated by steroids and lipid metabolites, and each member is responsible for a unique subset of genes for lipid and energy metabolism. PPAR activity is also regulated by post-translational modifications. In the Central Nervous System (CNS), PPARs are expressed in all types of nerve cells, and regulate many physiological processes, such as energy metabolism, redox homeostasis, autophagy, cell cycle and differentiation, etc. Following acute or chronic central nervous system injury, PPARs are involved in the regulation of various pathways including neuroinflammation, antioxidant responses, and survival/neurodegenerative mechanisms. To date, most studies have been directed to the gamma isoform, and very little to the effects of PPAR α and β/δ. It is rarely reported in the existing research whether PPAR γ is involved in the regulation of neuronal differentiation and whether PPAR γ is involved in the regulation of high-sugar-induced autophagy.

The inventor of the invention carries out a great deal of research to clarify the influence of hyperglycemia caused by gestational diabetes mellitus on the development of a fetal nervous system, and finds that hyperglycemia obviously induces the expression of an autophagy molecule Beclin-1 and inhibits the expression of p 62; and the expression level of PPAR γ is significantly increased. These results indicate that the high sugar environment induces autophagy in neural stem cell precursors, induces their death, and hinders their differentiation into neurons, and that PPAR γ is involved in this regulatory process. Further research shows that the PPAR gamma can effectively restore the levels of GFAP and Tuj1 and down-regulate the expression level of Beclin-1, so that the levels of P62 and LC3 II/LC 3I tend to be normal, and the regulation effect of the PPAR gamma on the autophagy activity and the neuron differentiation induced by high sugar is fully proved, namely the autophagy process can be remarkably inhibited by inhibiting the activity of the PPAR gamma, the neuron differentiation is promoted, and the occurrence of nervous diseases caused by high sugar is avoided.

Compared with the prior art, the invention has the following technical effects:

(1) the invention carries out deep research on the influence of gestational hyperglycemia on neuron differentiation, and discovers that hyperglycemia can remarkably induce the expression of autophagy molecules, inhibit the neuron differentiation and influence the normal development of a fetal nervous system;

(2) the invention defines a specific action mechanism of influence on neuron differentiation under a high-sugar condition mediated by PPAR gamma, namely the PPAR gamma expression level is obviously increased under the high-sugar environment, so that autophagy of neural stem cell precursor cells is induced, death of the neural stem cell precursor cells is induced, differentiation of the neural stem cell precursor cells to neurons is hindered, and further the development of a nervous system is realized;

(3) the invention discovers that the levels of GFAP and Tuj1 can be effectively recovered and the expression level of Beclin-1 is reduced through inhibiting the expression of PPAR gamma, so that the levels of P62 and LC3 II/LC 3I tend to be normal, the generation of a cell autophagy process is obviously inhibited, and the differentiation of neurons is promoted; discloses a new mechanism for effectively regulating neuron differentiation disorder caused by hyperglycemia by regulating the activity of a target PPAR gamma, and provides practical experimental evidence and scientific basis for clinically intervening and treating relevant neurological diseases caused by gestational diabetes.

Drawings

FIG. 1 is a diagram of the model of the differentiation of P19 mouse neural stem cell-like cells into neurons.

FIG. 2 is a schematic diagram showing the effect of high sugar on Tuj1 protein expression in neuronal stem cells.

FIG. 3 is a graph showing the quantitative results of the effect of high sugar on Tuj1 protein expression in neuronal stem cells.

FIG. 4 is a graph showing the results of quantitative analysis of the effect of high sugar on Tuj1 mRNA expression in neuronal stem cells.

FIG. 5 is a graph showing the effect of high sugar on GFAP protein expression in neuronal stem cells.

FIG. 6 is a graph showing the results of quantitative analysis of the effect of high sugar on GFAP protein expression in neuronal stem cells.

FIG. 7 is a graph showing the results of quantifying the effect of high sugar on GFAP mRNA expression in neuronal stem cells.

FIG. 8 is a schematic diagram showing the flow results of the effect of high sugar on apoptosis of neuronal stem cells.

FIG. 9 is a diagram showing the result of quantitative analysis of the effect of high sugar on apoptosis of neuronal stem cells.

FIG. 10 is a graph showing the effect of high sugar on the expression of biomarkers associated with autophagy.

FIG. 11 is a graph showing the results of quantitative analysis of the effect of high sugar on the expression of biomarkers associated with autophagy.

Fig. 12 is a graph showing the effect of shpara on PPAR γ protein expression in neuronal stem cells.

Fig. 13 is a diagram showing the results of quantitative analysis of the effect of shpara on PPAR γ protein expression in neuronal stem cells.

FIG. 14 is a graph showing the effect of shPPAR γ -1 on Tuj1 and GFAP protein expression in neuronal stem cells in a high-sugar environment.

Fig. 15 is a graph showing the results of quantitative analysis of the effect of shPPAR γ -1 on Tuj1 and GFAP protein expression in neuronal stem cells in a high-sugar environment.

Fig. 16 is a graph showing the effect of shpara γ -1 on cell autophagy-related biomarker expression in a high-sugar environment.

Fig. 17 is a diagram showing the results of quantitative analysis of the effect of shPPAR gamma-1 on cell autophagy-related biomarker expression in a high-sugar environment.

Fig. 18 is a graph showing the effect of shpara-1 on P19 cell differentiation in a high-sugar environment.

Fig. 19 is a graph showing the results of quantitative analysis of the effect of shpara-1 on P19 cell differentiation in a high-sugar environment.

Detailed Description

In order to make the objects, technical solutions and effects of the present invention clearer and clearer, the present invention is further described in detail below with reference to examples. It should be understood that the specific embodiments described herein are merely illustrative of the invention and are not intended to limit the invention.

Cell tools including P19 neural stem cell-like cells listed in the context of the present invention are commercially available and cultured according to conventional methods in cell biology, unless otherwise specified. All cell lines were identified by short tandem repeat analysis of the chinese typical culture collection (wuhan) and verified for the presence of mycoplasma contamination using a PCR assay kit (shanghai Biothrive Sci) while being cryopreserved in liquid nitrogen and used for subsequent experiments. The reagents used in the present invention are commercially available. The experimental methods and techniques used in the present invention, such as P19 cell culture, Western blot, molecular cloning, PCR, immunofluorescent staining, laser confocal, flow cytometry and animal experiments, are all conventional methods and techniques in the art.

Representative results from selection of biological experimental replicates are presented in the context figure, and data are presented as mean ± SD and mean ± SEM as specified in the figure. All experiments were repeated at least three times. Data were analyzed using GraphPad Prism 5.0 or SPSS 20.0 software. And comparing the difference of the mean values of two or more groups by using a t test or an analysis of variance. p < 0.05 was considered a significant difference.

Example 1 Effect of high sugar Environment on neuronal Stem cell differentiation

(1) Adding Retinoic Acid (RA) into P19 mouse neural stem cell-like cells to construct a model of differentiation of P19 cells into neurons (as shown in FIG. 1);

(2) dividing the cell models in the step (1) into two groups, wherein the group 1 is a control group, culturing is carried out under normal conditions (normal glucose, 5 mM), the group 2 is a high-sugar group, and high-sugar culture conditions (high-concentration glucose, 30 mM) are given, so that the gestational hyperglycemia is simulated, and the influence of high sugar on the differentiation of the neuron stem cells is observed;

(3) after the administration of high sugar, the mRNA and protein expression levels of Tuj1 and GFAP in the two groups of cells were measured by Western blot on

days

3, 5 and 7 after the induction of cell differentiation, respectively, and the cell growth was analyzed.

The results are shown in FIGS. 2-7. The results show that after high sugar treatment, the expression level of mRNA or protein of marker GFAP of both neuron-specific expression Tuj1 and glial neuron is obviously reduced in P19 cells under high sugar environment, and the difference is statistically significant. Thus proving that the high-sugar environment can generate obvious inhibition effect on the differentiation process of the neuron cells.

Further, analysis of P19 cells by flow cytometry revealed that the cell viability in the high-sugar group was significantly lower than that in the normal control group (see fig. 8-9), and the difference was statistically significant. Thus, the fact that the high sugar can remarkably induce the apoptosis of the neuron stem cells is proved.

Example 2 Effect of high sugars on autophagy function

(1) Adding Retinoic Acid (RA) into the neural stem cell-like cells of the P19 mouse to construct a differentiation model of the P19 cells into neurons;

(3) after administration of high sugar, protein expression levels of P62, Beclin-1 and PPAR γ were measured in the two groups of cells by Western blot on day 7 after induction of cell differentiation, respectively.

The results are shown in FIGS. 10 to 11. The results show that the levels of PPAR gamma and Beclin-1 in the neuron cells under the high-sugar treatment environment are obviously up-regulated, the level of P62 is obviously reduced, and the difference has statistical significance. During the autophagy process, the level of P62 is often closely related to autophagy activity, the increase of P62 level is generally considered as a sign of the inhibition of autophagy activity, and the decrease of P62 level indicates the occurrence of autophagy process. Beclin-1 is also an essential molecule in the formation process of autophagosome, and can mediate other autophagy proteins to localize in phagocytic vacuoles so as to regulate the formation and maturation of autophagosome, and the expression level of Beclin-1 is often increased remarkably in the autophagy process. From the above results, it is clear that the high sugar environment can significantly induce the expression of the autophagy molecule Beclin-1, while inhibiting the expression of P62. The above results together indicate that the high sugar environment can induce autophagy of neural stem cell precursor cells, induce death thereof, and hinder differentiation to neurons, and PPAR γ may be one of the key regulatory factors of the process.

Example 3 study of the modulatory effects of PPAR γ inhibitors on neuronal differentiation

(1) shRNA is designed and synthesized aiming at the mRNA sequence of the PPAR gamma, and is shPPAR gamma-1 (the sequence is shown in SEQ ID NO: 1), shPPAR gamma-2 (the sequence is shown in SEQ ID NO: 2) and shPPAR gamma-3 (the sequence is shown in SEQ ID NO: 3);

(2) an interference sequence designed for PPAR γ was then loaded onto plk.0 lentiviral vectors and gene sequencing identified whether the loading was successful. The P19 cells were then plated at 1X 10⁵Inoculating the culture medium into a 10cm culture dish at a density of one ml, and changing the culture medium into an OPTI-MEM culture medium on the next day;

(3) respectively taking shPPAR gamma-1, shPPAR gamma-2, shPPAR gamma-3 and shNC carrier to transfect according to the mass volume ratio of 1:2 of plasmid and transfection reagent, and replacing the culture medium to be a complete culture medium after 6 hours;

(4) proteins of P19 cells transfected with shPPAR γ -1, shPPAR γ -2, shPPAR γ -3 and shNC (empty vector) were extracted, and the expression level of the PPAR γ protein was examined using Western blot.

The results are shown in FIGS. 12-13. The results show that the activity of the intracellular PPAR gamma protein of the P19 cells transfected with the shPPAR gamma-1 and the shPPAR gamma-3 is remarkably inhibited, and the difference has statistical significance compared with the difference of an empty vector group, thereby proving that the shPPAR gamma-1 and the shPPAR gamma-3 can obviously inhibit the expression and the activity of the PPAR gamma.

Subsequently, the research on the effect of the shPPAR gamma on the neuron differentiation related protein under the high-sugar environment comprises the following specific steps:

(2) dividing the cell model of step (1) into four groups, wherein group 1 was a control group and cultured by normal conditions (normal glucose, 5 mM), group 2 was a high-sugar group and given high-sugar culture conditions (high-concentration glucose, 30 mM), group 3 was P19 cells transfected with an empty vector (shNC) and cultured by high-sugar conditions (high-concentration glucose, 30 mM), and group 4 was P19 cells transfected with shpara-1 and cultured by high-sugar conditions (high-concentration glucose, 30 mM);

(3) protein expression levels of Tuj1 and GFAP in both groups of cells were measured by Western blot 72h after induction of cell differentiation, respectively, after administration of high sugar.

The results are shown in FIGS. 14-15. The results show that under the condition of high sugar, the protein expression levels of Tuj1 specifically expressed by neurons in the cells of the group 2 and the group 3 and the marker GFAP of glial neurons are obviously inhibited; after the cells of the group 4 are transfected by the shPPAR gamma-1 to effectively inhibit the PPAR gamma, the expression activities of Tuj1 and GFAP are obviously up-regulated, even higher than that of a blank control group 1, wherein the level difference between the group 4 and the group 2 or the group 3 has statistical significance.

Further, the research on the effect of shPPAR gamma on autophagy function in a high-sugar environment comprises the following specific steps:

(3) protein expression levels of PPAR γ, Beclin-1, LC 3I, LC3 II and P62 in both groups of cells were examined by Western blot 72h after induction of cell differentiation, respectively, after administration of high sugar.

The results are shown in FIGS. 16-17. The results show that the levels of PPAR gamma, Beclin-1 and LC3 II in the cells of the

groups

2 and 3 are obviously up-regulated, the levels of LC 3I and P62 are obviously down-regulated, and the ratio of LC3 II/LC 3I is obviously increased, so that the autophagy degree in the cells of the

groups

2 and 3 can be obviously increased. In the cells of the group 4 transfected with the shPPAR γ, the levels of the PPAR γ, Beclin-1 and LC II are obviously reduced, the levels of the LC 3I and the P62 are obviously increased, and the ratio of the LC3 II to the LC 3I is obviously reduced and tends to be normal, even lower than that of the blank control group 1, so that the shPPAR γ can be definitely and remarkably inhibit the autophagy process in the cells.

Under the high-sugar treatment environment, the levels of PPAR gamma and Beclin-1 in the neuron cells are obviously up-regulated, the level of P62 is obviously reduced, and the difference has statistical significance.

Under the condition of high sugar, the protein expression levels of Tuj1 specifically expressed by neurons in the cells of the group 2 and the group 3 and a marker GFAP of glial neurons are obviously inhibited; after the transfection of the shPPAR gamma-1 to effectively inhibit the PPAR gamma, the expression activities of Tuj1 and GFAP are obviously up-regulated, even higher than that of a blank control group 1, wherein the level difference between a group 4 and a group 2 or a group 3 has statistical significance.

Furthermore, the research on the effect of the shPPAR gamma in the high-sugar environment on the neuron differentiation in the high-sugar environment comprises the following specific steps:

(1) p19 cells were cultured at 1X 10⁵The cells were inoculated in a 10cm dish and P19 cell-inducing solution was cultured in alpha MEM containing 5% fetal bovine serum and 0.5. mu.M trans-retinoic acid;

(2) changing the inducing liquid the next day and continuously culturing for four days to induce the differentiation of the P19 cells;

(3) on day four, the cell pellet was trypsinized and blown into single cells, and the cells were cultured using Neurobasal and B27 mature neuronal medium.

The results are shown in FIGS. 18-19. The results show that: under high sugar conditions, the percentage of P19 cells differentiating into positive neurons was reduced, while the percentage of P19 cells differentiating into positive neurons was restored after transfection of shpara-1 effectively inhibited PPAR γ. This suggests that high glucose inhibits differentiation of P19 cells into neurons, whereas inhibition of PPAR γ restores the potential of P19 cells to differentiate neurons.

In combination with the above results, the present invention clearly reveals the role and mechanism of PPAR γ in the process of neuronal differentiation in a high-sugar environment. The PPAR gamma has a regulation effect with autophagy activity and can mediate the autophagy process, so that the neuron differentiation is inhibited, and the normal development of a nervous system is influenced; after the interference of the PPAR gamma, the expression of the PPAR gamma in the cell can be obviously reduced, and the autophagy process of the cell is obviously inhibited, so that the differentiation of the neuron is promoted. Discloses a new mechanism of PPAR gamma for regulating and controlling neuron differentiation under a high-sugar environment, and provides practical experimental evidence and research direction for clinically intervening and treating relevant neurological diseases caused by gestational diabetes.

The above detailed description section specifically describes the analysis method according to the present invention. It should be noted that the above description is only for the purpose of helping those skilled in the art better understand the method and idea of the present invention, and not for the limitation of the related contents. The present invention may be appropriately adjusted or modified by those skilled in the art without departing from the principle of the present invention, and the adjustment and modification also fall within the scope of the present invention.

Sequence listing

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Claims

1. A pharmaceutical composition for treating neuronal differentiation disorders comprising a PPAR γ inhibitor selected from the group consisting of shpara designed based on the PPAR γ gene.

2. The pharmaceutical composition of claim 1, wherein the neuronal differentiation disorder is a diabetic neuronal differentiation disorder.

3. The pharmaceutical composition of claim 2, wherein the diabetic neuron differentiation disorder is a gestational diabetic neuron differentiation disorder.

4. The pharmaceutical composition of any one of claims 1-3, wherein the shpara is selected from one or more of shpara-1, shpara-2, shpara-3; the sequences of the shPPAR gamma-1, the shPPAR gamma-2 and the shPPAR gamma-3 are respectively shown as SEQ ID NO: 1. SEQ ID NO: 2. SEQ ID NO: 3, respectively.

Use of a PPAR γ inhibitor for the manufacture of a medicament for the treatment of a neuronal differentiation disorder.

6. The use according to claim 5, wherein the PPAR γ inhibitor is selected from the group consisting of shPPAR γ designed based on the PPAR γ gene.

7. The use of claim 6, wherein the shpara is selected from one or more of shpara-1, shpara-2, shpara-3; the sequences of the shPPAR gamma-1, the shPPAR gamma-2 and the shPPAR gamma-3 are respectively shown as SEQ ID NO: 1. SEQ ID NO: 2. SEQ ID NO: 3, respectively.

Use of a PPAR γ inhibitor in the manufacture of a medicament for promoting neuronal differentiation.

9. The use according to claim 8, wherein the PPAR γ inhibitor is selected from the group consisting of shpara designed based on the PPAR γ gene.

10. The use of claim 9, wherein the shpara is selected from one or more of shpara-1, shpara-2, shpara-3; the sequences of the shPPAR gamma-1, the shPPAR gamma-2 and the shPPAR gamma-3 are respectively shown as SEQ ID NO: 1. SEQ ID NO: 2. SEQ ID NO: 3, respectively.