Use of sodium butyrate in the manufacture of a medicament for treating an intracranial hemorrhage condition

Abstract

The application discloses application of sodium butyrate in preparation of a medicine for treating intracranial hemorrhage, in particular, application of sodium butyate in preparation of a medicine for treating neonatal intraventricular hemorrhage, researches show that the sodium butyate is used for preventive neuroprotection of IVH of high-risk premature infants (fetal age is less than 32 weeks), secondary brain injury is reduced, and long-term neurodevelopmental outcome is improved; the sodium butyate is used for treating neonatal intraventricular hemorrhage, brain ventricle expansion is reduced, neuron survival is increased, and nerve function is improved; and the sodium butyate can also improve motor function and cognitive function, and has the effect of comprehensive neuroprotection. The application provides a new medicine source for treating intracranial hemorrhage, in particular, neonatal intraventricular hemorrhage.

Description

Technical Field

[0001] This invention relates to the use of a drug for the prevention and treatment of hemorrhage, and particularly to the use of a drug for the prevention and treatment of intraventricular hemorrhage, belonging to the field of medicine. Background Technology

[0002] Sodium butyrate is the sodium salt of the short-chain fatty acid butyrate, primarily derived from the fermentation of dietary fiber by gut microbiota. In recent years, the protective effects of sodium butyrate on neurological diseases have received widespread attention. International research has shown that sodium butyrate possesses histone deacetylase inhibitor activity, exerting neuroprotective effects through epigenetic regulation. In adult models of stroke and traumatic brain injury, sodium butyrate can reduce neuronal apoptosis, inhibit neuroinflammatory responses, and promote neurological function recovery. Domestic research has explored the protective effects of sodium butyrate on neurodegenerative diseases, confirming its ability to improve cognitive impairment.

[0003] Intraventricular hemorrhage (IVH) is the most common type of intracranial hemorrhage in premature infants, especially those with a gestational age of less than 32 weeks and a birth weight of less than 1500 grams, with an incidence of approximately 15% to 20%. Severe IVH can lead to ventricular dilatation, hydrocephalus, and long-term neurodevelopmental disorders. Regarding the current state of research on IVH treatment, internationally, the main approaches are supportive care, cerebrospinal fluid drainage, and surgical interventions such as ventriculoperitoneal shunts, with a lack of effective neuroprotective drugs. In China, neonatology research on IVH prevention and treatment primarily focuses on perinatal management and coagulation function regulation, with relatively limited research on neuroprotective interventions after hemorrhage, and no approved specific neuroprotective drugs currently used clinically.

[0004] Currently, research on the application of sodium butyrate in neurological diseases mainly focuses on adult disease models, which has the following technical limitations: First, the applicable population is limited; existing studies have all focused on adult animals or adult patients, neglecting the specific population of newborns or premature infants, for whom there is a lack of effective neuroprotective drugs. Second, the disease model is limited; there is a lack of efficacy evaluations specifically for neonatal IVH. Third, the dosing regimen is unclear; neonates have significantly different drug metabolism characteristics compared to adults, lacking suitable dosage, route of administration, and treatment duration regimens for newborns, and the effective dose range in newborns is unknown. Fourth, developmental safety has not been assessed; the long-term effects on brain damage after neonatal IVH are unknown, and the potential impact of sodium butyrate on neonatal brain development is unclear, lacking developmental neuropharmacological data. Fifth, the mechanism of action is unclear; in the pathological context of neonatal IVH, the specific neuroprotective targets and molecular mechanisms of sodium butyrate have not been elucidated. Summary of the Invention

[0005] The purpose of this invention is to address the aforementioned technical deficiencies in existing treatments for neonatal intraventricular hemorrhage (IVH) by providing an application of sodium butyrate in the preparation of drugs for treating neonatal IVH. This invention applies sodium butyrate to the prevention and treatment of neonatal IVH, filling a gap in drug intervention in this field. It also clarifies the dosage, administration method, and treatment regimen for treating intraventricular hemorrhage. The invention evaluates the neuroprotective effects of sodium butyrate on neonatal IVH, including improvement of ventricular dilation, recovery of motor function, protection of cognitive function, and promotion of neuronal survival. Furthermore, it provides a complete chain of pharmacodynamic evidence, covering multiple levels including imaging, behavioral, and histological aspects.

[0006] To achieve the objectives of this invention, one aspect of this invention provides the use of sodium butyrate in the preparation of a medicament for treating intracranial hemorrhage.

[0007] The intracranial hemorrhage condition is intraventricular hemorrhage; preferably, it is neonatal intraventricular hemorrhage.

[0008] In particular, the newborns are premature infants with a gestational age of less than 32 weeks and a birth weight of less than 1500 grams.

[0009] In particular, the drug consists of sodium butyrate and a pharmaceutically acceptable carrier.

[0010] In particular, the purity of the sodium butyrate is ≥1%, preferably 95% or higher or 95-100% pure substance.

[0011] In particular, pharmaceutically acceptable carriers are generally recognized by healthcare professionals for use in this purpose and as inactive ingredients in pharmaceutical preparations. Compilations of pharmaceutically acceptable carriers can be found in reference books such as the *Handbook of Pharmaceutical Excipients* (2nd edition, edited by A. Wade and P. J. Weller; published by the American Pharmaceutical Association, Washington and The Pharmaceutical Press, London, 1994).

[0012] In particular, the drug exists in the form of tablets, capsules, pills, powders, granules, syrups, emulsions, etc.

[0013] In particular, the drug is preferably administered in the form of an oral solution or an enteric-coated formulation.

[0014] Another aspect of the present invention provides a medicament for treating intraventricular hemorrhage in newborns, wherein the medicament contains sodium butyrate.

[0015] In particular, the weight ratio of the sodium butyrate to the total weight of the drug or health product is 0.1:100-99:100.

[0016] In particular, the purity of the sodium butyrate is ≥1%, preferably 95% or higher or 95-100% pure substance.

[0017] The drug can be prepared into various dosage forms, such as tablets, capsules, pills, powders, granules, syrups, and solutions, using methods known in the art.

[0018] The present invention also provides a method for treating intraventricular hemorrhage in newborns, comprising administering a therapeutically effective dose of sodium butyrate to the newborn suffering from intraventricular hemorrhage, wherein the therapeutically effective dose is 1200-3600 mg / kg.d, preferably 3600 mg / kg.d.

[0019] Unless otherwise stated, the term "therapeutic effective dose" as used in this invention refers to the amount of drug required to produce an effective effect; the "therapeutic effective dose" is adjustable and variable, and is ultimately determined by medical personnel, taking into account factors such as the route of administration and the nature of the formulation, the recipient's weight, age and other general characteristics, as well as the nature and severity of the disease being treated.

[0020] This invention is the first to apply sodium butyrate to the prevention and treatment of neonatal intracranial hypothermia (IVH), filling a gap in drug intervention in this field. Secondly, it establishes an IVH model suitable for newborn rats and a sodium butyrate administration regimen, clarifying the optimal dose of 3600 mg / kg via gavage. Thirdly, it systematically evaluates the neuroprotective effects of sodium butyrate on neonatal IVH, including improvement in ventricular dilation, recovery of motor function, protection of cognitive function, and promotion of neuronal survival. Fourthly, it provides a complete chain of pharmacodynamic evidence, covering multiple levels including imaging, behavioral, and histological studies.

[0021] This invention relates to a novel use of sodium butyrate in the preparation of drugs for the prevention or treatment of neonatal intraventricular hemorrhage. The mechanism of action is as follows: after neonatal intraventricular hemorrhage, primary damage is caused by collagenase-induced vascular rupture and bleeding, while secondary damage includes hematoma compression, inflammatory response, neuronal apoptosis, and ventricular dilatation. Sodium butyrate intervention, through HDAC inhibition, leads to increased histone acetylation levels, which in turn upregulates the expression of neuroprotective genes, exerting anti-inflammatory, anti-apoptotic, and neurogenesis-promoting effects, ultimately achieving a protective effect of reduced ventricular dilatation, increased neuronal survival, and improved neurological function.

[0022] Sodium butyrate is used as a neuroprotective drug in the treatment of intraventricular hemorrhage in newborns.

[0023] The administration regimen is oral gavage (which can be developed into an oral formulation suitable for newborns), with a dosage of 3600 mg / kg (equivalent dose for newborn rats, which can be converted based on the body surface area of ​​newborns). The timing of administration is as early as possible after the onset of IVH, and the course of treatment is continued until 2 weeks after model establishment (which can be adjusted according to clinical conditions).

[0024] The present invention has the following advantages and benefits:

[0025] 1. This invention is the first to discover the neuroprotective effect of sodium butyrate on neonatal IVH, filling a gap in the field of drug intervention.

[0026] 2. The sodium butyrate of the present invention has multi-target protection for the treatment of neonatal IVH, and at the same time improves ventricular dilation, motor function, cognitive function and neuronal survival, thus having a comprehensive neuroprotective effect.

[0027] 3. The present invention has a convenient route of administration, a simple method of administration by gavage, and is easy to convert into an oral preparation suitable for newborns.

[0028] 4. Sodium butyrate is an endogenous short-chain fatty acid with good safety profile, making it suitable for newborns, a special population.

[0029] 5. The mechanism of sodium butyrate in treating IVH of the present invention is clear. It exerts an epigenetic regulatory effect through HDAC inhibition. The mechanism is clear and facilitates further in-depth research.

[0030] 6. There is currently a lack of effective drugs for neonatal IVH. The sodium butyrate of this invention has a clear therapeutic effect on neonatal IVH, a clear mechanism of action, and a clear need and prospect for clinical translation, with great potential for clinical translation.

[0031] 7. This invention also optimizes the dosage of sodium butyrate for treating neonatal IVH. Through experimental screening, 3600 mg / kg was determined to be the optimal dosage, providing a reference for clinical application. Attached Figure Description

[0032] Figure 1 The images show a comparison of intracranial magnetic resonance imaging (MRI) of newborn rats in each group 72 hours after modeling. The T2WI images show successful modeling and ventricular enlargement, which are used for model validation.

[0033] Figure 1A Comparison of ventricular area at the striatum and hippocampus levels in newborn rats 72 hours after modeling, including the control group, IVH group, and different NaB dose groups.

[0034] Figure 2 Comparative images of the cranial magnetic resonance imaging of newborn rats in different groups 2 weeks after sodium butyrate administration show that the ventricles of the sodium butyrate 3600 mg / kg group were significantly reduced, demonstrating that the core invention is that sodium butyrate improves ventricular dilation.

[0035] Figure 2A Comparison of ventricular area at the striatum and hippocampus levels in newborn rats two weeks after sodium butyrate administration, including the control group, IVH group, and different NaB dose groups.

[0036] Figure 3 A comparison chart showing the rotarod dwell time of rats in the IVH group, control group, and IVH plus NaB group;

[0037] Figure 4 Comparison chart of gripping test results for rats in the IVH group, control group, and IVH plus NaB group;

[0038] Figure 5 Heatmaps of rat trajectories in the Y-maze for the IVH group, control group, and IVH plus NaB group are shown. Figure 5 Figure A is a representative heatmap of the rat movement trajectory in the Y maze of the IVH group rats; Figure 5B is a representative heatmap of the rat movement trajectory in the Y maze of the control group rats; Figure 5C is a representative heatmap of the rat movement trajectory in the Y maze of the IVH+sodium butyrate group rats.

[0039] Figure 6 A comparison diagram of the spontaneous alternation rate in the Y maze of rats in the IVH group, control group, and IVH plus NaB group;

[0040] Figure 7 A comparison chart showing the total number of arm advances in the Y-maze for rats in the IVH group, control group, and IVH plus NaB group;

[0041] Figure 8 HE staining results for the IVH group, control group, and IVH plus NaB group show the structure of hippocampal and cortical tissues and neuronal arrangement, providing histological evidence of protection.

[0042] Figure 9 NeuN staining results for the IVH group, control group, and IVH plus NaB group show neuronal survival (fluorescence intensity) and provide evidence of neuronal protection.

[0043] Figure 10 The results of quantitative analysis of NeuN staining fluorescence intensity in the IVH group, control group, and IVH plus NaB group were presented. Quantitative statistical graphs showed the NeuN expression level in each brain region, providing quantitative pharmacodynamic evidence. Detailed Implementation

[0044] The present invention will be further described below with reference to specific embodiments, and the advantages and features of the present invention will become clearer as a result. However, these embodiments are merely exemplary and do not constitute any limitation on the scope of the present invention. Those skilled in the art should understand that modifications or substitutions can be made to the details and form of the technical solutions of the present invention without departing from the spirit and scope of the present invention, but all such modifications and substitutions fall within the protection scope of the present invention.

[0045] Experimental Example 1: Establishment of a neonatal rat model of intraventricular hemorrhage

[0046] 1. Test materials

[0047] 1) Laboratory animals

[0048] The experimental animals were newborn SD rats that were 5 days old, which is equivalent to human premature infants with a gestational age of 28 to 32 weeks.

[0049] 2) Experimental reagents

[0050] Sodium butyrate, collagenase VII, isoflurane, and artificial cerebrospinal fluid.

[0051] 3) Experimental equipment

[0052] Stereostat, microinjection pump, 3.0T small animal magnetic resonance imaging system, rotator, gripping force measuring instrument and Y maze.

[0053] 2. Test methods

[0054] Newborn SD rats aged 5 days (P5) were placed in an anesthesia induction box and induced with 3% to 5% isoflurane. After the righting reflex disappeared, the concentration was maintained at 1.5%. The rats were placed in a prone position and fixed to the operating table with a stereotaxic device. The height of the incisor bar was adjusted to -3.3 mm, and the ear bar was fixed.

[0055] Routine scalp preparation and disinfection, including iodine disinfection of the scalp.

[0056] Make a longitudinal incision of about 10 mm along the midline of the scalp to expose the skull, the anterior fontanelle, and the sagittal suture. Fix the glass electrode to the stereotaxic instrument with the tip of the needle positioned in the right striatum (1 mm from the beak of the anterior fontanelle, 4 mm lateral to the midline, 3.5 mm deep; coordinates: AP -1.0 mm, ML ±4.0 mm, DV -3.5 mm). Adjust the stereotaxic parameters to position the injection needle 1.0 mm posterior to the anterior fontanelle (-1.0 mm anteroposterior diameter), 4.0 mm to the right of the midline (+4.0 mm medial and lateral), and 3.5 mm subdurally (-3.5 mm dorsoventrally). After adjusting the microinjection pump parameters, use a 10 μL microsyringe to inject 2 μL of collagenase VII (dissolved in artificial cerebrospinal fluid, 0.5 units per μL, injection rate 1 μL / min) into the right striatum near the ventricle. Leave the needle in place for 10 min to allow the collagenase to fully diffuse into the brain region and prevent backflow. Then, slowly lift the microsyringe (0.5 mm / min) and suture the scalp incision. After the surgery, place the animal on a constant temperature heating pad (set to 35 °C) and monitor it continuously until it fully recovers from anesthesia before returning it to its mother's cage.

[0057] The sham surgery group received an equal volume of artificial cerebrospinal fluid instead of collagenase injected into the right striatum near the ventricle, serving as a control group.

[0058] At 72 hours after modeling, the area of ​​the rat ventricles was detected by MRI-T2WI. After the rats were anesthetized and fixed, T2WI sequence scanning was performed, and the outline of the ventricles was delineated and the area was calculated using ImageJ software.

[0059] MRI-T2WI 72 hours after modeling showed that the model was successfully established in all collagenase-induced hemorrhage groups, and there was no statistically significant difference in ventricular area between groups (P > 0.05), indicating that the model was stable.

[0060] Experimental Example 2: Study on the neuroprotective effect of sodium butyrate on intraventricular hemorrhage in neonatal rats

[0061] (1) Grouping and administration

[0062] Following the method in Experiment 1, collagenase was used to induce intraventricular hemorrhage in newborn rats, and a sham-operated group was set up as the control group.

[0063] Sodium butyrate was administered via gavage immediately after modeling, with a treatment course of 2 weeks.

[0064] Sodium butyrate was dissolved in sterile physiological saline and freshly prepared at concentrations of 0, 60, 120, and 360 mg / mL; the dosage was 0, 600, 1200, and 3600 mg / kg (i.e., 10 mL / kg volume); the administration route was by gavage using a blunt-tipped gavage needle; the administration time was immediately after model establishment, once daily for 14 consecutive days.

[0065] Based on the dosage of sodium butyrate, the patients were divided into five concentration gradient groups: no administration, 600 mg / kg, 1200 mg / kg, 2400 mg / kg, and 3600 mg / kg. The patients were given the medication by gavage once a day for 14 consecutive days.

[0066] The control group (i.e., the sham surgery group) and the model group (i.e., the IVH group) were given the same volume of normal saline by gavage; the sodium butyrate administration group was given different doses of sodium butyrate by gavage, and the sham surgery group and the IVH group were given the same volume of normal saline by gavage.

[0067] (2) Cranial magnetic resonance imaging examination

[0068] All newborn rats underwent T2WI cranial magnetic resonance imaging (MRI) at 72 hours and 2 weeks after modeling.

[0069] The results showed that 72 hours after modeling, all collagenase-induced hemorrhage groups were successfully modeled, with intraventricular hemorrhage and ventricular dilation occurring, while the sham surgery group showed no hemorrhage or ventricular dilation.

[0070] There were no significant differences in the degree of ventricular dilation at the striatal and hippocampal levels between the IVH group (i.e., the model group) and different sodium butyrate dosage groups before sodium butyrate treatment (e.g., ...). Figure 1 , Figure 1A ).

[0071] Two weeks after sodium butyrate treatment, a follow-up cranial MRI was performed. MRI-T2WI was used to assess the degree of ventricular dilation. The results are as follows:

[0072] Compared to the IVH group, the sodium butyrate administration groups (1200-3600 mg / kg) showed a significant reduction in ventricular size at the striatal and hippocampal levels (P < 0.05), confirming that sodium butyrate can significantly improve ventricular dilation after hemorrhage (e.g., Figure 2 , Figure 2A Therefore, sodium butyrate at 3600 mg / kg was chosen as the subsequent experimental dose.

[0073] (3) Improvement of motor and cognitive outcomes in rats after intraventricular hemorrhage:

[0074] To further evaluate the long-term neuroprotective effects of sodium butyrate at the behavioral level, we administered sodium butyrate at 3600 mg / kg by gavage for 2 weeks after model establishment. At 3-4 weeks after birth (equivalent to human infancy), we used the rotarod test to assess motor coordination, the grip strength test to assess muscle strength and motor function, and the Y-maze test to assess spatial working memory.

[0075] Behavioral tests are conducted using single-blind or double-blind methods.

[0076] The test results of the rotatable bar test, grip strength measurement, and Y-maze test are as follows: Figure 3-7 .

[0077] ①Rotating rod experiment

[0078] Rats were pre-trained for 3 days. The formal test used an accelerated mode (4 to 40 revolutions per minute for 300 seconds), and the fall latency was recorded. The test results are as follows: Figure 3 The results showed that the average dwell time in the IVH group was significantly shorter than that in the control group, decreasing to 52% of that in the control group, indicating motor coordination disorder; after sodium butyrate gavage, the dwell time in the rotarod test was significantly improved, recovering to 70% of that in the control group, and the improvement rate increased by 18%.

[0079] ② Grip test

[0080] With the rats' hind limbs and trunk fixed, only their forelimbs grasped the grip bar. The bar was pulled horizontally backward, and the maximum pulling force was recorded. This was repeated 5 times, and the maximum value was recorded. The test results are as follows: Figure 4 The results showed that the maximum grip strength of the forelimbs in the IVH group decreased significantly, down to 74% of that in the control group, indicating motor coordination disorder; after oral administration of sodium butyrate, the maximum grip strength of the forelimbs improved significantly, recovering to 90% of that in the control group, with an improvement rate of 16%.

[0081] ③Y-maze experiment

[0082] Rats were allowed to adapt to the Y-maze for 5 minutes, followed by a formal test lasting 8 minutes. The order and number of arm movements were recorded, and the spontaneous alternation rate was calculated. The results were examined as follows: Figure 5-7 ,in Figure 5 This is a representative heatmap of the rat's movement trajectory in the Y-maze. Figure 5 Figure A shows a representative heatmap of the rat movement trajectory in the Y-maze of the IVH group; Figure B shows a representative heatmap of the rat movement trajectory in the Y-maze of the control group; Figure C shows a representative heatmap of the rat movement trajectory in the Y-maze of the IVH + sodium butyrate group.

[0083] like Figure 6 The spontaneous alternation rate in the Y maze was significantly reduced in the IVH group, dropping to 60% of the control group, indicating a decline in spatial working memory. After oral administration of sodium butyrate, the spontaneous alternation rate recovered to 85% of the control group, with an improvement rate of 25 percentage points.

[0084] The results of the total number of arm advances by rats in the Y-maze are as follows: Figure 7 The total number of arm insertions was significantly reduced in the IVH group, dropping to 74% of the control group, indicating a decline in spatial working memory. After sodium butyrate gavage, the total number of arm insertions recovered to 78% of the control group.

[0085] Based on the above behavioral indicators, oral administration of sodium butyrate can improve motor coordination and cognitive memory in IVH rats, providing reliable behavioral evidence for neuronal protection at the pathological level.

[0086] (4) Reduce neuronal damage after intraventricular hemorrhage:

[0087] To verify the protective effect of sodium butyrate on neuronal survival after IVH at the histological level, brain tissue was collected simultaneously at the MRI endpoint (i.e., 2 weeks after modeling) for coronal brain slices with HE staining to detect brain tissue pathology; NeuN immunofluorescence staining was used to detect neuronal survival. The steps for NeuN immunofluorescence quantification were: perfusion brain harvesting, frozen sectioning, immunofluorescence staining (NeuN primary antibody plus fluorescent secondary antibody), confocal microscopy imaging, and ImageJ quantitative fluorescence intensity.

[0088] HE test results are as follows Figure 8 NeuN test results are as follows Figure 9 Imaging analysis was conducted using single-blind or double-blind methods.

[0089] Figure 8 HE pathology findings: In the IVH group, the hippocampus and cortex volume decreased, and the pyramidal neurons were arranged in a disordered manner; while in the sodium butyrate administration group (i.e., IVH plus NaB group), the hippocampus and cortex volume increased, the neurons were arranged in a regular manner, and the neuronal damage was improved.

[0090] Figure 9 The NeuN detection results showed that the fluorescence intensity of NeuN in the hippocampus and cortex of the IVH group was significantly reduced (P < 0.05), indicating neuronal damage or maturation disorders; while the fluorescence intensity of NeuN in the CA1 region, DG region and cortex of the IVH+NaB group treated with sodium butyrate was significantly increased, indicating improved neuronal damage; the fluorescence intensity was significantly higher than that of the IVH group (P < 0.05), the density of NeuN-positive neurons rebounded, and the cell bodies were full.

[0091] Quantitative analysis of NeuN staining fluorescence intensity, such as Figure 10 The analysis results show that:

[0092] The fluorescence intensity of NeuN in the hippocampus and cortex of the IVH group was significantly reduced (P < 0.05), suggesting neuronal damage or maturation disorders in the cortex and hippocampus of the IVH group; after administration of sodium butyrate, the fluorescence intensity of NeuN in the CA1 and DG regions of the hippocampus and the cortex was significantly increased, suggesting improvement in neuronal damage.

[0093] Based on the above experimental results, it can be seen that oral administration of sodium butyrate at 3600 mg / kg can significantly improve ventricular dilation after neonatal IVH, effectively improve motor coordination function (rotary rod, grasp test), significantly improve spatial working memory ability (Y maze), promote neuronal survival, and reduce pathological damage to brain tissue.

[0094] Example 1: Neuroprotective effect of sodium butyrate on intraventricular hemorrhage in neonatal rats

[0095] 1. Establish an IVH model

[0096] Eight to ten animals were used in each group. P5 newborn rats were placed in an anesthesia induction box and induced with 3% to 5% isoflurane. After the righting reflex disappeared, the concentration was maintained at 1.5%. The rats were fixed in a prone position on a stereotaxic apparatus, with the incisor bar height adjusted to -3.3 mm and the ear bar fixed. The scalp was disinfected with iodine, and a longitudinal incision of about 10 mm was made along the midline to expose the anterior fontanelle and sagittal suture. The stereotaxic apparatus was adjusted so that the injection needle was positioned 1.0 mm posterior to the anterior fontanelle (-1.0 mm anteroposterior diameter) and 4.0 mm to the right of the midline. (4.0 mm on the inner and outer sides), 3.5 mm subdurally (3.5 mm negative on the dorsal and ventral sides); use a 10 μL microsyringe to draw 2 μL of collagenase VII solution (0.5 units per μL, dissolved in artificial cerebrospinal fluid), connect it to a microinjection pump, and inject at a rate of 1 μL per minute. Leave the needle in place for 10 minutes after injection; slowly withdraw the needle (0.5 mm per minute) and suture the scalp incision; place the mouse on a constant temperature heating pad (35 degrees Celsius) and monitor continuously until it is fully awake, then return it to the mother mouse cage.

[0097] The sodium butyrate administration regimen was as follows: sodium butyrate was dissolved in sterile saline and freshly prepared to a concentration of 360 mg / mL; the dosage was 3600 mg / kg (i.e., 10 mL / kg volume); the administration route was by gavage using a blunt-tipped gavage needle; the administration time was immediately after model establishment, once daily for 14 consecutive days; the control treatments were the IVH group and the Sham group, which received the same volume of saline by gavage.

[0098] 2. Experimental Methods

[0099] MRI scans were performed 72 hours after modeling (to validate the model) and 14 days after drug administration (to evaluate efficacy). The parameters were T2WI sequence (TR / TE=2500 / 36 ms, slice thickness 0.5 mm, no gap). The ventricular contours were manually delineated at the hippocampal level and the ventricular area was calculated using ImageJ software.

[0100] Behavioral testing should be conducted 3 to 4 weeks after birth, using the following methods:

[0101] 1) Rotating bar experiment

[0102] The test is conducted 3-4 weeks after birth (equivalent to early childhood in humans), including the rotata test (pre-training for 3 days, 4 rotatas per minute, 300 seconds, for 3 consecutive days; the formal test uses an accelerated mode of 4 to 40 rotatas per minute, and the fall latency is recorded, and the average value is taken for 3 consecutive tests).

[0103] 2) Grip force measurement

[0104] With the hind limbs and trunk fixed, only the forelimbs grasp the grip bar, and the maximum pulling force is recorded by pulling horizontally backward. The maximum value is recorded after 5 consecutive tests.

[0105] 3) Y-maze experiment

[0106] The three arms are all 40 cm x 8 cm x 15 cm in length, with an included angle of 120 degrees. After 5 minutes of adaptation, conduct a formal 8-minute test, record the order of arm entry and calculate the spontaneous alternation rate. The formula is: the number of correct alternations divided by the total number of arm entrys minus 2, then multiplied by 100%.

[0107] Histological examination, specifically including:

[0108] Specimen preparation (heart perfusion with 4% paraformaldehyde fixative, followed by rinsing with physiological saline and then perfusion with fixative; complete removal of brain tissue followed by fixation for 24 hours; coronal sections 30 μm thick, 1 out of every 5 sections), HE staining (sections mounted on slides, hematoxylin-eosin staining, dehydrated and cleared, mounted, observed under a light microscope for hippocampal CA1, CA3 and cortical structures), and NeuN immunofluorescence staining (blocking with 5% normal goat serum and 0.3% Triton X-100 at room temperature for 1 hour; primary antibody incubation with rabbit anti-NeuN antibody 1:500, overnight at 4°C; secondary antibody incubation with Alexa Fluor 488-labeled goat anti-rabbit IgG 1:1000, at room temperature for 2 hours; counterstaining of cell nuclei with DAPI and mounting; observation under a confocal microscope (Zeiss LSM 880, 20x objective); quantitative analysis of fluorescence intensity using ImageJ software, calculating the average fluorescence intensity per field of view).

[0109] 3. Experimental Results

[0110] The survival rate of newborn rats in the sodium butyrate administration group was greater than 90%, no obvious adverse reactions were observed in the sodium butyrate administration group, and the weight gain was not significantly different from that in the control group.

[0111] Regarding MRI results, right ventricular enlargement was observed in both the IVH group and the IVH plus NaB group 72 hours after modeling, with no significant difference in ventricular area (P > 0.05), confirming successful modeling and good consistency. 14 days after drug administration, the ventricular area of ​​the IVH group continued to expand, while the ventricular area of ​​the sodium butyrate 3600mg / kg group was 40-50% smaller than that of the IVH group (P < 0.05).

[0112] In terms of behavioral results, the mean rotarod time was 161.5 seconds in the control group, 83.6 seconds in the IVH group, and 112.2 seconds in the IVH plus NaB group. The difference between the IVH group and the control group was P < 0.001, and the difference between the IVH plus NaB group and the IVH group was P < 0.05.

[0113] The average grip strength was 478.7 g in the control group, 355.6 g in the IVH group, and 429.2 g in the IVH plus NaB group. The difference between the IVH group and the control group was P < 0.01, and the difference between the IVH plus NaB group and the IVH group was P < 0.05.

[0114] Y-maze results: The spontaneous alternation rate was 31.7% in the control group, 18.9% in the IVH group, and 26.9% in the IVH plus NaB group. The difference between the IVH group and the control group was P < 0.01, and the difference between the IVH plus NaB group and the IVH group was P < 0.05.

[0115] In terms of histological results, HE staining showed that the pyramidal neurons in the CA1 region of the hippocampus in the IVH group were sparsely and disordered, and the cortical thickness was thinner. In the sodium butyrate group, the neurons were arranged more regularly, and the cortical thickness was increased. NeuN staining showed that the number of NeuN-positive cells in the CA1, DG and cortical regions of the hippocampus in the IVH group was reduced, and the fluorescence intensity was decreased. In the sodium butyrate group, the NeuN fluorescence intensity in each brain region was significantly increased (P < 0.05).

[0116] The conclusion of this embodiment is that oral administration of sodium butyrate at 3600 mg / kg can significantly improve ventricular dilation, motor dysfunction, cognitive impairment and neuronal damage in neonatal rats after intraventricular hemorrhage, and has a good neuroprotective effect, providing experimental evidence for the use of sodium butyrate in the prevention and treatment of neonatal intraventricular hemorrhage.

[0117] The above embodiments of the present invention are merely exemplary and do not constitute any limitation on the scope of the present invention. Those skilled in the art should understand that modifications or substitutions can be made to the details and form of the technical solutions of the present invention without departing from the spirit and scope thereof, but all such modifications and substitutions fall within the protection scope of the present invention.

Claims

1. The use of sodium butyrate in the preparation of a drug for treating intracranial hemorrhage.

2. The application as described in claim 1, characterized in that, The drug consists of sodium butyrate and a pharmaceutically acceptable carrier.

3. The application as described in claim 1 or 2, characterized in that, The drug exists in the form of tablets, capsules, pills, powders, granules, syrups, or emulsions.

4. The application as described in claim 1 or 2, characterized in that, The purity of the sodium butyrate is ≥1%.

5. A drug for treating intraventricular hemorrhage in newborns, characterized in that, It contains sodium butyrate.

6. The drug as described in claim 5, characterized in that, The weight ratio of the sodium butyrate to the total weight of the drug or health product is (0.1-99):

100.

7. The drug as described in claim 5 or 6, characterized in that, The purity of the sodium butyrate is ≥1%.

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