Novel application of transcription factor FOXD1

Abstract

The invention relates to novel application of a transcription factor FOXD1, in particular to application of the transcription factor FOXD1 in regulating pituitary tumor disease. On the basis of a gene chip method, 10 pituitary tumor case samples and 5 normal comparison samples are sequenced, and a backup gene is screened. In addition, QPCR verification and siRNA interference experiment are performed, results show that the FOXD1 is highly related to pituitary tumor, and proliferation of pituitary tumor cells is slowed down by interfering FOXD 1 gene expression. The results provide a novel idea for clinical diagnosis and novel treatment of the pituitary tumor.

Description

technical field

[0001] The invention relates to the field of biomedicine, in particular to the new use of transcription factor FOXD1, and more specifically to the role of transcription factor FOXD1 in regulating pituitary tumor diseases. Background technique

[0002] Most of the regulation of eukaryotic transcriptional level is achieved through complex interactions between cis-regulatory elements and trans-regulatory factors. The cis-regulatory element is the site of regulation, and the trans-regulatory factors that interact with the cis-regulatory element as a regulatory tool are proteins that can directly or indirectly recognize the cis-regulatory element and participate in the regulation of transcription. General trans-regulatory factors are also called transcription factors (Transcription Factor, TF). In a broad sense, trans-regulators also include other proteins such as RNA polymerase. Transcription factors are a class of proteins with specific functions. By binding t...

Examples

Embodiment 1

[0035] Example 1 Differential expression of FOXD1 gene

[0036] 1. Collect samples

[0037] Patients with pituitary tumors: Among the 10 patients with pituitary tumors, there were 5 males, aged 28-73, with an average age of 47; 5 females, aged 25-68, with an average age of 44. Inclusion criteria: patients underwent transsphenoidal pituitary tumor resection, brain MRI and other imaging data were available before surgery, and postoperative pathological sections confirmed pituitary tumor. With the consent of the hospital ethics committee, the materials were collected after the patients signed the informed consent.

[0038] Normal pituitary gland tissue: 5 cases were taken from the Anatomy Department of the University, and endocrine-related diseases were excluded. The sampling method was the same as that of patients with pituitary tumors.

[0039] The required tissues were taken during the operation, washed with normal saline, immediately put into cryopreservation tubes treated...

Embodiment 2

[0050] Example 2 qPCR experiment to verify the differential expression of FOXD1 gene

[0051] 1. Collect samples

[0052] According to the standard of Example 1, 40 patients with pituitary tumors were collected, and their pituitary tumor tissues were collected. 30 normal pituitary tissues were collected.

[0053] 2. RNA extraction and quality analysis

[0054] use Reagent (invitrogen, Cat. No. 15596-018) was used for sample RNA extraction.

[0055] The specific operation is as follows:

[0056] Collect the samples and freeze them in liquid nitrogen. After taking them out, put the tissue into a pre-cooled mortar for grinding. After the tissue sample is powdered:

[0057] ① Add Trizol and store at room temperature for 5 minutes;

[0058] ② Add 0.2ml of chloroform, vibrate the centrifuge tube vigorously, mix well, and place it at room temperature for 5-10 minutes;

[0059] ③ After 15 minutes of high-speed centrifugation at 12000rpm, suck the upper aqueous phase (absorb 70...

Embodiment 3

[0085] Example 3 Inhibition of FOXD1 Gene Expression

[0086] 1. siRNA design and synthesis

[0087]

[0088]

[0089] The above siRNA sequences and negative control siRNA sequences (NC-siRNA) were provided by Shanghai Gemma Pharmaceutical Technology Co., Ltd.:

[0090] 2. Culture and transfection of pituitary tumor cells

[0091] 2.1 Cell culture

[0092] The pituitary tumor tissue pieces were immersed in RPMI-1640 medium, washed 3 times with DPBS, cut into pieces, digested with 0.25% trypsin at room temperature for 1 hour, and added an equal volume of RPMI-1640 medium containing 10% FCS to stop digestion, adding a final concentration of 0.87 %NH 4 Cl acted at 37°C for 5mim; filtered through a 70-mesh cell sieve to remove tissue debris, centrifuged at 1000r / min for 5min, and treated with RPMI- The cell pellet was resuspended in 1640 medium, and cultured in a carbon dioxide cell incubator with 5% carbon dioxide, 37°C and saturated humidity. The medium was changed on...

technical field

[0001] The invention relates to the field of biomedicine, in particular to the new use of transcription factor FOXD1, and more specifically to the role of transcription factor FOXD1 in regulating pituitary tumor diseases. Background technique

[0002] Most of the regulation of eukaryotic transcriptional level is achieved through complex interactions between cis-regulatory elements and trans-regulatory factors. The cis-regulatory element is the site of regulation, and the trans-regulatory factors that interact with the cis-regulatory element as a regulatory tool are proteins that can directly or indirectly recognize the cis-regulatory element and participate in the regulation of transcription. General trans-regulatory factors are also called transcription factors (Transcription Factor, TF). In a broad sense, trans-regulators also include other proteins such as RNA polymerase. Transcription factors are a class of proteins with specific functions. By binding t...

Examples

Embodiment 1

[0035] Example 1 Differential expression of FOXD1 gene

[0036] 1. Collect samples

[0037] Patients with pituitary tumors: Among the 10 patients with pituitary tumors, there were 5 males, aged 28-73, with an average age of 47; 5 females, aged 25-68, with an average age of 44. Inclusion criteria: patients underwent transsphenoidal pituitary tumor resection, brain MRI and other imaging data were available before surgery, and postoperative pathological sections confirmed pituitary tumor. With the consent of the hospital ethics committee, the materials were collected after the patients signed the informed consent.

[0038] Normal pituitary gland tissue: 5 cases were taken from the Anatomy Department of the University, and endocrine-related diseases were excluded. The sampling method was the same as that of patients with pituitary tumors.

[0039] The required tissues were taken during the operation, washed with normal saline, immediately put into cryopreservation tubes treated...

Embodiment 2

[0050] Example 2 qPCR experiment to verify the differential expression of FOXD1 gene

[0051] 1. Collect samples

[0052] According to the standard of Example 1, 40 patients with pituitary tumors were collected, and their pituitary tumor tissues were collected. 30 normal pituitary tissues were collected.

[0053] 2. RNA extraction and quality analysis

[0054] use Reagent (invitrogen, Cat. No. 15596-018) was used for sample RNA extraction.

[0055] The specific operation is as follows:

[0056] Collect the samples and freeze them in liquid nitrogen. After taking them out, put the tissue into a pre-cooled mortar for grinding. After the tissue sample is powdered:

[0057] ① Add Trizol and store at room temperature for 5 minutes;

[0058] ② Add 0.2ml of chloroform, vibrate the centrifuge tube vigorously, mix well, and place it at room temperature for 5-10 minutes;

[0059] ③ After 15 minutes of high-speed centrifugation at 12000rpm, suck the upper aqueous phase (absorb 70...

Embodiment 3

[0085] Example 3 Inhibition of FOXD1 Gene Expression

[0086] 1. siRNA design and synthesis

[0087]

[0088]

[0089] The above siRNA sequences and negative control siRNA sequences (NC-siRNA) were provided by Shanghai Gemma Pharmaceutical Technology Co., Ltd.:

[0090] 2. Culture and transfection of pituitary tumor cells

[0091] 2.1 Cell culture

[0092] The pituitary tumor tissue pieces were immersed in RPMI-1640 medium, washed 3 times with DPBS, cut into pieces, digested with 0.25% trypsin at room temperature for 1 hour, and added an equal volume of RPMI-1640 medium containing 10% FCS to stop digestion, adding a final concentration of 0.87 %NH 4 Cl acted at 37°C for 5mim; filtered through a 70-mesh cell sieve to remove tissue debris, centrifuged at 1000r / min for 5min, and treated with RPMI- The cell pellet was resuspended in 1640 medium, and cultured in a carbon dioxide cell incubator with 5% carbon dioxide, 37°C and saturated humidity. The medium was changed on...

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