1.Quality changes of volatile oil and chlorogenic acid compounds during extraction process of Artemisiae Argyi Folium: process analysis based on chemical composition, physicochemical properties, and biological activity.
Dan-Dan YANG ; Hao-Zhou HUANG ; Xin-Ming CHEN ; Lin HUANG ; Ya-Nan HE ; Zhen-Feng WU ; Xiao-Ming BAO ; Ding-Kun ZHANG ; Ming YANG
China Journal of Chinese Materia Medica 2025;50(11):3001-3012
To explore the variation laws of volatile oil during the extraction process of Artemisiae Argyi Folium and its impact on the quality of the medicinal solution, as well as to achieve precise control of the extraction process, this study employed headspace solid phase microextraction gas chromatography-mass spectrometry(HS-SPME-GC-MS) in combination with multiple light scattering techniques to conduct a comprehensive analysis, identification, and characterization of the changes in volatile components and the physical properties of the medicinal solution during the extraction process. A total of 82 volatile compounds were identified using the HS-SPME-GC-MS technique, including 21 alcohols, 15 alkenes, 14 ketones, 9 acids, 6 aldehydes, 5 phenols, 3 esters, and 9 other types of compounds. At different extraction time points(15, 30, 45, and 60 min), 71, 72, 64, and 44 compounds were identified in the medicinal solution, respectively. It was observed that the content of volatile components gradually decreased with the extension of extraction time. Through multivariate statistical analysis, four compounds with significant differences during different extraction time intervals were identified, namely 1,8-cineole, terpinen-4-ol, 3-octanone, and camphor. RESULTS:: from multiple light scattering techniques indicated that at 15 minutes of extraction, the transmittance of the medicinal solution was the lowest(25%), the particle size was the largest(0.325-0.350 nm), and the stability index(turbiscan stability index, TSI) was the highest(0-2.5). With the extension of extraction time, the light transmittance of the medicinal solution improved, stability was enhanced, and the particle size decreased. These laws of physicochemical property changes provide important basis for the control of Artemisiae Argyi Folium extraction process. In addition, the changes in the bioactivity of Artemisiae Argyi Folium extracts during the extraction process were investigated through mouse writhing tests and antimicrobial assays. The results indicated that the analgesic and antimicrobial effects of the medicinal solution were strongest at the 15-minute extracting point. In summary, the findings of this study demonstrate that the content of volatile oil in Artemisiae Argyi Folium extracts gradually decreases with the extension of extraction time, and the variation in volatile oil content directly influences the physicochemical properties and pharmacological efficacy of the medicinal solution. This discovery provides important scientific reference for the optimization of Artemisiae Argyi Folium extraction processes and the development and application of process analytical technologies.
Oils, Volatile/pharmacology*
;
Artemisia/chemistry*
;
Gas Chromatography-Mass Spectrometry
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Drugs, Chinese Herbal/pharmacology*
;
Chlorogenic Acid/pharmacology*
;
Solid Phase Microextraction
;
Quality Control
2.Clinical observation on the efficacy of self-made anatomical tower-shaped pad combined with splint plaster fixation in the treatment of unstable the 2nd to 5th metacarpal fractures.
Xiao-Bao WU ; Xiao-Rong ZHENG ; Zhen-Sheng WANG ; Zhi-Qiang LIN
China Journal of Orthopaedics and Traumatology 2025;38(7):747-751
OBJECTIVE:
To evaluate the clinical efficacy of self-made anatomical tower-shaped pads combined with splint plaster fixation in the treatment of unstable 2nd to 5th metacarpal fractures and to provide a reference for clinical practice.
METHODS:
A retrospective analysis was conducted on 98 patients with unstable 2nd to 5th metacarpal fractures treated with self-made anatomical tower-shaped pads combined with splint plaster fixation between January 2019 and December 2022. There were 74 males and 24 females, aged from 19 to 63 years old with an average of (41.58±7.23) years old. The total active movement (TAM) score was used to evaluate the metacarpophalangeal joint function. Complications and fracture healing time were recorded.
RESULTS:
All patients were followed up for a period ranging from 1 to 5 months, with an average duration of (3.45±1.03) months. Among the 98 patients, anatomical alignment was achieved in 75 patients, with 68 patients still maintaining anatomical alignment after fixation removal. Functional alignment was achieved in 23 patients, with 20 patients maintaining functional alignment even after fixation removal. There were 10 patients with displacement after reduction. The average fracture healing time was (5.78±1.14) weeks, and the fracture healed well without any angular or rotational deformities. TAM score was utilized to assess the hand function of patients 3 months post-treatment. The extension range of motion (10.72±1.35)° and flexion range of motion (83.19±4.08)° of the metacarpophalangeal joint were significantly higher than the pre-treatment values of (8.25±0.68)° and (70.35±2.36)°, respectively. These differences were statistically significant (P<0.05). TAM outcomes were excellent in 92 cases, good in 5, fair in 1, and poor in none.
CONCLUSION
The implementation of self-made anatomical tower-shaped pad combined with splint plaster fixation is effective for the conservative management of unstable 2nd to 5th metacarpal fractures.
Humans
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Male
;
Female
;
Adult
;
Middle Aged
;
Metacarpal Bones/surgery*
;
Splints
;
Retrospective Studies
;
Fractures, Bone/therapy*
;
Casts, Surgical
;
Young Adult
;
Fracture Fixation, Internal/instrumentation*
3.GOLM1 promotes cholesterol gallstone formation via ABCG5-mediated cholesterol efflux in metabolic dysfunction-associated steatohepatitis livers
Yi-Tong LI ; Wei-Qing SHAO ; Zhen-Mei CHEN ; Xiao-Chen MA ; Chen-He YI ; Bao-Rui TAO ; Bo ZHANG ; Yue MA ; Guo ZHANG ; Rui ZHANG ; Yan GENG ; Jing LIN ; Jin-Hong CHEN
Clinical and Molecular Hepatology 2025;31(2):409-425
Background/Aims:
Metabolic dysfunction-associated steatohepatitis (MASH) is a significant risk factor for gallstone formation, but mechanisms underlying MASH-related gallstone formation remain unclear. Golgi membrane protein 1 (GOLM1) participates in hepatic cholesterol metabolism and is upregulated in MASH. Here, we aimed to explore the role of GOLM1 in MASH-related gallstone formation.
Methods:
The UK Biobank cohort was used for etiological analysis. GOLM1 knockout (GOLM1-/-) and wild-type (WT) mice were fed with a high-fat diet (HFD). Livers were excised for histology and immunohistochemistry analysis. Gallbladders were collected to calculate incidence of cholesterol gallstones (CGSs). Biles were collected for biliary lipid analysis. HepG2 cells were used to explore underlying mechanisms. Human liver samples were used for clinical validation.
Results:
MASH patients had a greater risk of cholelithiasis. All HFD-fed mice developed MASH, and the incidence of gallstones was 16.7% and 75.0% in GOLM1-/- and WT mice, respectively. GOLM1-/- decreased biliary cholesterol concentration and output. In vivo and in vitro assays confirmed that GOLM1 facilitated cholesterol efflux through upregulating ATP binding cassette transporter subfamily G member 5 (ABCG5). Mechanistically, GOLM1 translocated into nucleus to promote osteopontin (OPN) transcription, thus stimulating ABCG5-mediated cholesterol efflux. Moreover, GOLM1 was upregulated by interleukin-1β (IL-1β) in a dose-dependent manner. Finally, we confirmed that IL-1β, GOLM1, OPN, and ABCG5 were enhanced in livers of MASH patients with CGSs.
Conclusions
In MASH livers, upregulation of GOLM1 by IL-1β increases ABCG5-mediated cholesterol efflux in an OPN-dependent manner, promoting CGS formation. GOLM1 has the potential to be a molecular hub interconnecting MASH and CGSs.
4.GOLM1 promotes cholesterol gallstone formation via ABCG5-mediated cholesterol efflux in metabolic dysfunction-associated steatohepatitis livers
Yi-Tong LI ; Wei-Qing SHAO ; Zhen-Mei CHEN ; Xiao-Chen MA ; Chen-He YI ; Bao-Rui TAO ; Bo ZHANG ; Yue MA ; Guo ZHANG ; Rui ZHANG ; Yan GENG ; Jing LIN ; Jin-Hong CHEN
Clinical and Molecular Hepatology 2025;31(2):409-425
Background/Aims:
Metabolic dysfunction-associated steatohepatitis (MASH) is a significant risk factor for gallstone formation, but mechanisms underlying MASH-related gallstone formation remain unclear. Golgi membrane protein 1 (GOLM1) participates in hepatic cholesterol metabolism and is upregulated in MASH. Here, we aimed to explore the role of GOLM1 in MASH-related gallstone formation.
Methods:
The UK Biobank cohort was used for etiological analysis. GOLM1 knockout (GOLM1-/-) and wild-type (WT) mice were fed with a high-fat diet (HFD). Livers were excised for histology and immunohistochemistry analysis. Gallbladders were collected to calculate incidence of cholesterol gallstones (CGSs). Biles were collected for biliary lipid analysis. HepG2 cells were used to explore underlying mechanisms. Human liver samples were used for clinical validation.
Results:
MASH patients had a greater risk of cholelithiasis. All HFD-fed mice developed MASH, and the incidence of gallstones was 16.7% and 75.0% in GOLM1-/- and WT mice, respectively. GOLM1-/- decreased biliary cholesterol concentration and output. In vivo and in vitro assays confirmed that GOLM1 facilitated cholesterol efflux through upregulating ATP binding cassette transporter subfamily G member 5 (ABCG5). Mechanistically, GOLM1 translocated into nucleus to promote osteopontin (OPN) transcription, thus stimulating ABCG5-mediated cholesterol efflux. Moreover, GOLM1 was upregulated by interleukin-1β (IL-1β) in a dose-dependent manner. Finally, we confirmed that IL-1β, GOLM1, OPN, and ABCG5 were enhanced in livers of MASH patients with CGSs.
Conclusions
In MASH livers, upregulation of GOLM1 by IL-1β increases ABCG5-mediated cholesterol efflux in an OPN-dependent manner, promoting CGS formation. GOLM1 has the potential to be a molecular hub interconnecting MASH and CGSs.
5.Enzyme-directed Immobilization Strategies for Biosensor Applications
Xing-Bao WANG ; Yao-Hong MA ; Yun-Long XUE ; Xiao-Zhen HUANG ; Yue SHAO ; Yi YU ; Bing-Lian WANG ; Qing-Ai LIU ; Li-He ZHANG ; Wei-Li GONG
Progress in Biochemistry and Biophysics 2025;52(2):374-394
Immobilized enzyme-based enzyme electrode biosensors, characterized by high sensitivity and efficiency, strong specificity, and compact size, demonstrate broad application prospects in life science research, disease diagnosis and monitoring, etc. Immobilization of enzyme is a critical step in determining the performance (stability, sensitivity, and reproducibility) of the biosensors. Random immobilization (physical adsorption, covalent cross-linking, etc.) can easily bring about problems, such as decreased enzyme activity and relatively unstable immobilization. Whereas, directional immobilization utilizing amino acid residue mutation, affinity peptide fusion, or nucleotide-specific binding to restrict the orientation of the enzymes provides new possibilities to solve the problems caused by random immobilization. In this paper, the principles, advantages and disadvantages and the application progress of enzyme electrode biosensors of different directional immobilization strategies for enzyme molecular sensing elements by specific amino acids (lysine, histidine, cysteine, unnatural amino acid) with functional groups introduced based on site-specific mutation, affinity peptides (gold binding peptides, carbon binding peptides, carbohydrate binding domains) fused through genetic engineering, and specific binding between nucleotides and target enzymes (proteins) were reviewed, and the application fields, advantages and limitations of various immobilized enzyme interface characterization techniques were discussed, hoping to provide theoretical and technical guidance for the creation of high-performance enzyme sensing elements and the manufacture of enzyme electrode sensors.
6.GOLM1 promotes cholesterol gallstone formation via ABCG5-mediated cholesterol efflux in metabolic dysfunction-associated steatohepatitis livers
Yi-Tong LI ; Wei-Qing SHAO ; Zhen-Mei CHEN ; Xiao-Chen MA ; Chen-He YI ; Bao-Rui TAO ; Bo ZHANG ; Yue MA ; Guo ZHANG ; Rui ZHANG ; Yan GENG ; Jing LIN ; Jin-Hong CHEN
Clinical and Molecular Hepatology 2025;31(2):409-425
Background/Aims:
Metabolic dysfunction-associated steatohepatitis (MASH) is a significant risk factor for gallstone formation, but mechanisms underlying MASH-related gallstone formation remain unclear. Golgi membrane protein 1 (GOLM1) participates in hepatic cholesterol metabolism and is upregulated in MASH. Here, we aimed to explore the role of GOLM1 in MASH-related gallstone formation.
Methods:
The UK Biobank cohort was used for etiological analysis. GOLM1 knockout (GOLM1-/-) and wild-type (WT) mice were fed with a high-fat diet (HFD). Livers were excised for histology and immunohistochemistry analysis. Gallbladders were collected to calculate incidence of cholesterol gallstones (CGSs). Biles were collected for biliary lipid analysis. HepG2 cells were used to explore underlying mechanisms. Human liver samples were used for clinical validation.
Results:
MASH patients had a greater risk of cholelithiasis. All HFD-fed mice developed MASH, and the incidence of gallstones was 16.7% and 75.0% in GOLM1-/- and WT mice, respectively. GOLM1-/- decreased biliary cholesterol concentration and output. In vivo and in vitro assays confirmed that GOLM1 facilitated cholesterol efflux through upregulating ATP binding cassette transporter subfamily G member 5 (ABCG5). Mechanistically, GOLM1 translocated into nucleus to promote osteopontin (OPN) transcription, thus stimulating ABCG5-mediated cholesterol efflux. Moreover, GOLM1 was upregulated by interleukin-1β (IL-1β) in a dose-dependent manner. Finally, we confirmed that IL-1β, GOLM1, OPN, and ABCG5 were enhanced in livers of MASH patients with CGSs.
Conclusions
In MASH livers, upregulation of GOLM1 by IL-1β increases ABCG5-mediated cholesterol efflux in an OPN-dependent manner, promoting CGS formation. GOLM1 has the potential to be a molecular hub interconnecting MASH and CGSs.
7.Analysis of completion rate of tumor evaluation at initial assessment and after neoadjuvant therapy for mid and low rectal cancer : a national multicenter real-world study
Kexuan LI ; Tixian XIAO ; Xiaodong WANG ; Bin WU ; Guole LIN ; Yuchen GUO ; Ming QU ; Si WU ; Xiaodong YANG ; Yinshengbo′er BAO ; Baohua WANG ; Fan ZHANG ; Xiangwang YU ; Beizhan NIU ; Junyang LU ; Lai XU ; Guannan ZHANG ; Zhen SUN ; Guoyou ZHANG ; Yan SHI ; Hong JIANG ; Yongjing TIAN ; Yongxiang LI ; Hongwei YAO ; Jun XUE ; Quan WANG ; Lie YANG ; Qian LIU ; Yi XIAO
Chinese Journal of Digestive Surgery 2025;24(1):113-119
Objective:To investigate the completion rate of tumor evaluation at initial assessment and after neoadjuvant therapy for mid and low rectal cancer patients in the national multicenter real-world database.Methods:The prospective real-world study was conducted. The clinicopathological data of 1 074 patients who underwent surgical treatment for mid and low rectal cancer in 47 national medical institutions, including Peking Union Medical College Hospital et al, from May 12,2023 to May 11,2024 were collected. Observation indicators: (1) clinical characteristics of patients with mid and low rectal cancer; (2) initial colonoscopy and pathologic evaluation of tumors in patients with mid and low rectal cancer; (3) initial imaging evaluation of patients with mid and low rectal cancer; (4) imaging evaluation after neoadjuvant therapy for patients with mid and low rectal cancer. Measurement data with normal distribution were represented as Mean± SD, and measurement data with skewed distribution were represented as M( Q1, Q3). Count data were described as absoluter numbers and/or percentages. Results:(1) Clinical characteristics of patients with mid and low rectal cancer. Of the 1 074 patients, there were 713 males and 361 females, aged 63(56,70)years. The body mass index of 1 074 patients was 24(21,26)kg/m 2.For American Society of Anesthesiologists classification, there were 147 cases of stage Ⅰ, 641 cases of stage Ⅱ, 157 cases of stage Ⅲ, 2 cases of stage Ⅳ, and there were 127 cases missing data. (2) Initial colonoscopy and pathologic evaluation of tumors in patients with mid and low rectal cancer. Of the 1 074 patients, there were 787 cases (73.28%) undergoing complete colonoscopy, and there were only 197 cases (18.34%) undergoing immunohistochemical evaluation of all four mismatch repair proteins. (3) Initial imaging evaluation of patients with mid and low rectal cancer. Of the 1 074 patients, there were 842(78.40%) patients completing magnetic resonance imaging (MRI) or ultrasound evaluation, and there were 914(85.10%) patients completing chest, abdomen, and pelvis enhanced computed tomography (CT) evaluation. In the 149 patients completing rectal ultrasound evaluation, there were 122 cases (81.88%) comple-ting T staging evaluation, and there were 81 cases (54.36%) completing N staging evaluation. In the 808 patients completing rectal MRI evaluation, there were 708 cases (87.62%) completing T staging evaluation, and there were 590 cases (73.02%) completing N staging evaluation. (4) Imaging evalua-tion after neoadjuvant therapy for patients with mid and low rectal cancer. Of the 388 patients with neoadjuvant therapy, there were 332 patients (85.57%) completing MRI or ultrasound evaluation, and there were 327 patients (84.28%) completing chest, abdomen, and pelvis enhanced CT evalua-tion. In the 70 patients completing rectal ultrasound evaluation, there were 65 cases (92.86%) com-pleting T staging evaluation, and there were 49 cases (70.00%) completing N staging evaluation. In the 327 patients completing rectal MRI evaluation, there were 246 cases (75.23%) completing T staging, and there were 228 cases (69.72%) completing N staging evaluation. Conclusion:The com-pletion rate of tumor imaging evaluation at initial assessment and after neoadjuvant therapy for mid and low rectal cancer patients on a national scale is relatively good.
8.Analysis of completion rate of tumor evaluation at initial assessment and after neoadjuvant therapy for mid and low rectal cancer : a national multicenter real-world study
Kexuan LI ; Tixian XIAO ; Xiaodong WANG ; Bin WU ; Guole LIN ; Yuchen GUO ; Ming QU ; Si WU ; Xiaodong YANG ; Yinshengbo′er BAO ; Baohua WANG ; Fan ZHANG ; Xiangwang YU ; Beizhan NIU ; Junyang LU ; Lai XU ; Guannan ZHANG ; Zhen SUN ; Guoyou ZHANG ; Yan SHI ; Hong JIANG ; Yongjing TIAN ; Yongxiang LI ; Hongwei YAO ; Jun XUE ; Quan WANG ; Lie YANG ; Qian LIU ; Yi XIAO
Chinese Journal of Digestive Surgery 2025;24(1):113-119
Objective:To investigate the completion rate of tumor evaluation at initial assessment and after neoadjuvant therapy for mid and low rectal cancer patients in the national multicenter real-world database.Methods:The prospective real-world study was conducted. The clinicopathological data of 1 074 patients who underwent surgical treatment for mid and low rectal cancer in 47 national medical institutions, including Peking Union Medical College Hospital et al, from May 12,2023 to May 11,2024 were collected. Observation indicators: (1) clinical characteristics of patients with mid and low rectal cancer; (2) initial colonoscopy and pathologic evaluation of tumors in patients with mid and low rectal cancer; (3) initial imaging evaluation of patients with mid and low rectal cancer; (4) imaging evaluation after neoadjuvant therapy for patients with mid and low rectal cancer. Measurement data with normal distribution were represented as Mean± SD, and measurement data with skewed distribution were represented as M( Q1, Q3). Count data were described as absoluter numbers and/or percentages. Results:(1) Clinical characteristics of patients with mid and low rectal cancer. Of the 1 074 patients, there were 713 males and 361 females, aged 63(56,70)years. The body mass index of 1 074 patients was 24(21,26)kg/m 2.For American Society of Anesthesiologists classification, there were 147 cases of stage Ⅰ, 641 cases of stage Ⅱ, 157 cases of stage Ⅲ, 2 cases of stage Ⅳ, and there were 127 cases missing data. (2) Initial colonoscopy and pathologic evaluation of tumors in patients with mid and low rectal cancer. Of the 1 074 patients, there were 787 cases (73.28%) undergoing complete colonoscopy, and there were only 197 cases (18.34%) undergoing immunohistochemical evaluation of all four mismatch repair proteins. (3) Initial imaging evaluation of patients with mid and low rectal cancer. Of the 1 074 patients, there were 842(78.40%) patients completing magnetic resonance imaging (MRI) or ultrasound evaluation, and there were 914(85.10%) patients completing chest, abdomen, and pelvis enhanced computed tomography (CT) evaluation. In the 149 patients completing rectal ultrasound evaluation, there were 122 cases (81.88%) comple-ting T staging evaluation, and there were 81 cases (54.36%) completing N staging evaluation. In the 808 patients completing rectal MRI evaluation, there were 708 cases (87.62%) completing T staging evaluation, and there were 590 cases (73.02%) completing N staging evaluation. (4) Imaging evalua-tion after neoadjuvant therapy for patients with mid and low rectal cancer. Of the 388 patients with neoadjuvant therapy, there were 332 patients (85.57%) completing MRI or ultrasound evaluation, and there were 327 patients (84.28%) completing chest, abdomen, and pelvis enhanced CT evalua-tion. In the 70 patients completing rectal ultrasound evaluation, there were 65 cases (92.86%) com-pleting T staging evaluation, and there were 49 cases (70.00%) completing N staging evaluation. In the 327 patients completing rectal MRI evaluation, there were 246 cases (75.23%) completing T staging, and there were 228 cases (69.72%) completing N staging evaluation. Conclusion:The com-pletion rate of tumor imaging evaluation at initial assessment and after neoadjuvant therapy for mid and low rectal cancer patients on a national scale is relatively good.
9.Finite element analysis of titanium rods after vertebral column decancellation osteotomy for ankylosing spondylitis
Bao-Ke SU ; Yong-Qing WANG ; Zhi-Jie KANG ; Hai-Yan WANG ; Feng JIN ; Xiao-He LI ; Zhen-Hua CAO ; Jia-Xuan HUO ; Yong ZHU ; Feng LI
Acta Anatomica Sinica 2024;55(3):339-344
Objective To analyze the stress changes of thoracic vertebra(T)11-sacrum(S)titanium rods in patients with ankylosing spondylitis after vertebral column decancellation(VCD)osteotomy,and provide reference for the selection and improvement of titanium rods before surgery.Methods The original data of the continuous scanning tomographic images of patients with ankylosing spondylitis after VCD osteotomy were imported into Mimics 21.0 in DICOM format,and T11-S vertebrae,screws and titanium rods were respectively reconstructed.They were imported into 3-Matic to establish a preliminary geometric modeling,and then processed with noise removal,paving,smoothing,etc.The improved model was imported into Hypermesh 10 software for grid division,and the material was imported into ANSYS 19.2 to display the finite element model after attribute assignment,Set the boundary and load conditions,and measure the stress value at the connection between the screw and the titanium rod.Results Under neutral position,forward bending,lateral bending,and axial rotation conditions,the titanium rod had the highest stress at the upper vertebrae(T11)and the lowest stress at the top vertebrae(L3);Under the backward extension condition,the titanium rod has the highest stress at the lower end vertebra(L5).Conclusion In the upper and lower vertebrae,it is possible to consider increasing the diameter of the titanium rod,enhancing its hardness,or changing it to a double rod.
10.Standardized operational protocol for the China Human Brain Bank Consortium(2nd edition)
Xue WANG ; Zhen CHEN ; Juan-Li WU ; Nai-Li WANG ; Di ZHANG ; Juan DU ; Liang YU ; Wan-Ru DUAN ; Peng-Hao LIU ; Han-Lin ZHANG ; Can HUANG ; Yue-Shan PIAO ; Ke-Qing ZHU ; Ai-Min BAO ; Jing ZHANG ; Yi SHEN ; Chao MA ; Wen-Ying QIU ; Xiao-Jing QIAN
Acta Anatomica Sinica 2024;55(6):734-745
Human brain banks use a standardized protocol to collect,process and store post-mortem human brains and related tissues,along with relevant clinical information,and to provide the tissue samples and data as a resource to foster neuroscience research according to a standardized operating protocols(SOP).Human brain bank serves as the foundation for neuroscience research and the diagnosis of neurological disorders,highlighting the crucial rule of ensuring the consistency of standardized quality for brain tissue samples.The first version of SOP in 2017 was published by the China Human Brain Bank Consortium.As members increases from different regions in China,a revised SOP was drafted by experts from the China Human Brain Bank Consortium to meet the growing demands for neuroscience research.The revised SOP places a strong emphasis on ethical standards,incorporates neuropathological evaluation of brain regions,and provides clarity on spinal cord sampling and pathological assessment.Notable enhancements in this updated version of the SOP include reinforced ethical guidelines,inclusion of matching controls in recruitment,and expansion of brain regions to be sampled for neuropathological evaluation.

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