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28 July 2023, Volume 43 Issue 7 Previous Issue   
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Biomaterials and regenerative medicine column
Innovative research team achievement column
Basic research
Clinical research
Evidence-based medicine
Review


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Biomaterials and regenerative medicine column

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Construction of acellular cartilage matrix/silk fibroin scaffold and its cartilage tissue engineering study WANG Qianyi, RAN Xinyue, ZHANG Peiling, CI Zheng, LEI Dong, ZHOU Guangdong 2023, 43 (7):  795-803.  doi: 10.3969/j.issn.1674-8115.2023.07.001 Abstract ( 419 )   HTML ( 27 )   PDF (4286KB) ( 554 )   Objective ·To construct a bioactivity tissue engineering scaffold with double network cross-linking for cartilage tissue regeneration using an acellular cartilage matrix (ACM) with a natural silk fibroin (SF) biomaterial. Methods ·The cell-associated immunogenic components were removed by nuclease digestion, and the extracellular matrix-associated glycoproteins and collagen structures were retained, The efficiency of cartilage tissue decellularization was measured by spectrophotometry by using DNA, histoglycosaminoglycan and collagen quantification kits. ACM and SF were configured into a mixed solution, and the nucleophilic cross-linking reaction with the hydroxyl and carboxyl groups contained in both was carried out by adding ethylene glycol diglycidyl ether. Then it was freeze-dried to make porous bionic scaffolds (n=5). At the same time, porous scaffolds containing only ACM or SF were prepared by the same method (n=5). The microstructure of the scaffolds was observed by scanning electron microscopy (SEM), and the mechanical strength, elastic modulus and resilience of different groups of scaffolds were evaluated by mechanical tests. The internal and external nutrient exchange capacity of the scaffolds was reacted by water absorption rate. Chondrocytes from rabbit ears were isolated, cultured, and seeded on ACM-SF scaffolds. After 1, 4, and 7 days of culture, the adhesion, distribution, and matrix secretion of the cells on the scaffolds were observed by SEM, and the viability status of the cells was determined by double-staining of live and dead cells. CCK-8 method was used to determine the cytotoxicity of the scaffolds. The cells were implanted subcutaneously in nude mice, cultured in vivo for 4 and 8 weeks, and finally removed for histological testing. Differences between groups were tested by One-Way ANOVA. Statistical significance was accepted at a value of P<0.05. Results ·After enzymatic digestion, almost no cells remained in the acellular matrix, and the active components of the extracellular matrix were retained. The composite scaffold prepared by ACM-SF has interconnected microporous structure and good elasticity, and could recover its original shape after repeated compression in the wet state. The water absorption rate of ACM-SF reached nearly 20 times, which provided an effective material exchange condition for the cell adhesion environment. Histological tests showed that the ACM-SF scaffold regenerated homogeneous, typical cartilage tissue in vivo. Conclusion ·ACM-SF composite porous scaffold has a good bionic microenvironment and can be applied to tissue engineering cartilage regeneration. Figures and Tables | References | Related Articles | Metrics

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Effect of hydrogel stiffness on nucleus pulposus cell phenotypes in vitro and its repairment of intervertebral disc in vivo CHEN Zehao, LÜ Zhendong, ZHANG Zhen, CUI Wenguo, ZHANG Yuhui 2023, 43 (7):  804-813.  doi: 10.3969/j.issn.1674-8115.2023.07.002 Abstract ( 463 )   HTML ( 34 )   PDF (4157KB) ( 321 )   Objective ·To investigate the effect of hydrogel stiffness on nucleus pulposus cell phenotype and its function in repairing intervertebral disc degeneration in rats. Methods ·Methacrylate gelatin (GelMA) hydrogels with different concentrations were constructed. The stiffness of the hydrogels was investigated by using rheological analysis and uniaxial compression test. The microstructure and morphology of the hydrogels were observed by scanning electron microscopy (SEM). Nucleus pulposus cells with normal phenotype were inoculated on the surface of GelMA hydrogels. The biocompatibility of the hydrogel was evaluated by live-dead cell staining and the growth pattern of nucleus pulposus cells on hydrogels with different stiffness was observed with phalloidin staining under microscope. Immunofluorescence staining was performed to examine the nuclear localization of Yes-associated protein (YAP) and real-time quantitative reverse transcription PCR (qRT-PCR) was used to detect the expression levels of nucleus pulposus cell-associated genes [neural cell adhesion molecule 1 (Ncam-1), aggrecan (Acan), sex-determing region of Y chromosome (SRY)-box transcription factor 9 (Sox9)]. A rat caudal acupuncture intervertebral disc degeneration model was established. Nucleus pulposus cells cultured on different hydrogels were harvested and injected into the degenerated discs separately. Four weeks after surgery, magnetic resonance imaging (MRI) was performed to analyze the water content of the intervertebral discs in each group. Histological tests were performed to examine the disc structure and proteoglycan levels. Results ·The elastic modulus of the hydrogels was 1 kPa and 200 kPa when the concentration of GelMA prepolymerisation solution was at 4% and 15% respectively. SEM observation revealed that the hydrogels showed a loose and porous microstructure, and the porosity of hydrogels decreased significantly with the decrease of their stiffness. In vitro experiments demonstrated that both GelMA hydrogel mediums showed good biocompatibility and the ability to support cell proliferation. Nucleus pulposus cells cultured on the soft matrix (4%GelMA) had a lower elongation and spreading area than those cultured on the stiff matrix (15%GelMA), showing a tendency of YAP concentration in the cytoplasm. The gene expression of nucleus pulposus cells was examined and the levels of Sox9, Acan and Ncam-1 in the soft matrix hydrogel group were 23.7, 6.6 and 12.7 times of those in the control group respectively. In vivo experiments on rat disc degeneration showed that the soft hydrogel matrix group had higher disc water content and structural integrity than the stiff hydrogel matrix group. Conclusion ·Compared to stiff GelMA hydrogels, hydrogels with low stiffness better maintain the growth phenotypes in the nucleus pulposus cells and have better therapeutic effect on disc degeneration in vivo. Figures and Tables | References | Related Articles | Metrics

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Feasibility study on the preparation of artificial small blood vessel by fluorinated decellularized rabbit aorta ZHANG Rujie, ZHOU Guangdong, WANG Jianbo, WENG Guangdong, LEI Dong, GONG Wenhui 2023, 43 (7):  814-820.  doi: 10.3969/j.issn.1674-8115.2023.07.003 Abstract ( 163 )   HTML ( 13 )   PDF (5019KB) ( 209 )   Objective ·To explore the feasibility of fluorinated decellularized rabbit aorta as a small artificial blood vessel for tissue engineering. Methods ·The obtained rabbit aorta was decellularized in combination with Triton X-100, sodium deoxycholate (SD), sodium dodecyl sulfate (SDS), DNAse, and RNAse. Hematoxylin-eosin staining (H-E staining), Masson staining and Verhoff-von Gieson staining were performed in the decellularized group and undecellularized group, respectively. The effect of decellularization was identified by field emission scanning electron microscope, and the morphological changes of decellularized blood vessels were observed. The decellularized rabbit aorta was used as the arterialized artificial small vessel scaffold, and the decellularized small vessel intima was modified with liquid perfluorocarbons coating to prepare a new type of artificial small vessel. The characteristic groups of the artificial small vessel were qualitatively and quantitatively determined. The dissipation time of liquid on the inner surface of the vessel and the flow of liquid on the surface of the vessel tilted at 45° were observed to analyze the hydrophobicity of the vessel. The blood vessels in the decellularized group and the fluorinated group were implanted with platelet-rich plasma, incubated, and observed under an electron microscope to evaluate the antiplatelet aggregation in vitro. The balloon pressure pump was connected to the aorta of the undecellularized group, decellularized group and fluoride group for bursting pressure test. Results ·Histological observation of blood vessels showed that the combination could effectively remove cells while retaining collagen and elastic fibers, and there was no damage to the intima under the electron microscope. There was no significant difference in the pressure blasting test among the three groups. In the hydrophobicity experiment, the retention time of water droplets on the membrane of the fluorinated group was over 5 min, and no obvious water marks were left on the 45° inclined plate. In the platelet adhesion test, intimal aggregation activated platelets in the decellularized group, while they were inhibited in the fluorinated group. Conclusion ·The decellularized blood vessels have good mechanical properties and physical stability by combined decellularization, and the fluorinated coating makes the blood vessels have good anticoagulant and biocompatibility. Figures and Tables | References | Related Articles | Metrics

Innovative research team achievement column

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Screening of AAK1 interaction proteins and its role in regulating global translation level in cells JIANG Guixian, HU Ronggui, WU Hao 2023, 43 (7):  821-828.  doi: 10.3969/j.issn.1674-8115.2023.07.004 Abstract ( 313 )   HTML ( 27 )   PDF (2709KB) ( 392 )   Objective ·To investigate noval interacting partners for adaptor-associated protein kinase 1 (AAK1) and AAK1-mediated biological functions besides clathrin-mediated endocytosis. Methods ·The labeled AAK1 vector and the blank control vector were transfected in HEK-293T cells, and the potential AAK1 interacting proteins were obtained by co-immunoprecipitation with agar-specific gel and mass spectrometry. Further verifications were performed by CoIP and fluorescence-based imaging. Recombinant proteins were purified in vitro and the direct interaction between proteins were confirmed by glutathione-S-transferase pulldown (GST Pulldown) assay. The regulation of AAK1 in the global protein synthesis was explored by puromycin incorporation assay. Results ·Mass spectrometry results showed that AAK1 was associated with a series of proteins, including fragile X mental retardation syndrome-related protein 1 (FXR1), FXR2 and fragile X mental retardation protein 1 (FMRP). Enriching with anti-FLAG agarose gels after exogenous transfecting of AAK1-3xFLAG and FMRP-MYC plasmids, the expression of FMRP-MYC was detected. The expression of FMRP could also be detected by CoIP with endogenous AAK1 antibodies. Fluorescence-based imaging showed that they were spatially colocalized in the cytoplasm. GST Pulldown assay showed that FMRP could pulldown recombinant HIS6-AAK1 protein. Puromycin incorporation assay showed that in the same amount of time, the number of newly synthesized peptides labeled with puromycin was positively correlated with AAK1 protein expression. Conclusion ·AAK1 directly interacts with FMRP in cytoplasm and could up-regulate global protein synthesis level. Figures and Tables | References | Related Articles | Metrics

Basic research

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Mechanism of blood-brain barrier damage caused by the inhibition of Wnt7/β-catenin pathway induced by endoplasmic reticulum stress in cerebrovascular endothelial cells after stroke DONG Haiping, XIE Haiyi, MA Xiaoxiao, WANG Zhenhong 2023, 43 (7):  829-838.  doi: 10.3969/j.issn.1674-8115.2023.07.005 Abstract ( 389 )   HTML ( 23 )   PDF (3501KB) ( 417 )   Objective ·To investigate whether the inactivation of Wnt7/β-catenin signaling causes the destruction of the blood-brain barrier (BBB) in cerebrovascular endothelial cells after ischemic stroke, and investigate whether endoplasmic reticulum stress bursts mediates the inhibition of Wnt7/β-catenin pathway. Methods ·The model of middle cerebral artery occlusion (MCAO) in mice was established by a monofilament nylon suture with a round tip, which was used to temporarily occlude the middle cerebral artery for 60 min. MCAO model mice were intraperitoneally injected with the endoplasmic reticulum stress blocker 4-phenylbutyric acid (4-PBA) as 4-PBA+MCAO group. Sham surgery group (Sham group) was set. Twenty-four hours after MCAO, Evans blue (EB) was used to measure the BBB permeability. The brain water content was calculated by dry-wet weight ratio, and the adhesion of cerebrovascular endothelial cells and pericytes in mice was measured by immunofluorescence. Human brain microvascular endothelial cells (HBMECs) were used to establish an oxygen and glucose deprivation (OGD) model for 4 h, and then cultured with 4-PBA for 24 h. Cells were divided into blank control group, OGD group, and OGD+4-PBA group for CCK-8 assay to determine the cell viability. FITC-labeled bovine serum albumin (FITC-BSA) was used to detect the cell permeability. The secretion of platelet-derived growth factor β (PDGF-β) was measured by ELISA. Fluo-3 AM fluorescence probe was used to detect the fluorescence intensity of cells to assess intracellular Ca2+ concentration, and reactive oxygen species (ROS) content was measured by CM-H2DCFDA fluorescence probe to clarify the endoplasmic reticulum stress state. Western blotting was used to examine the expression of connexins, including zonula occludens-1 (ZO-1) and claudin-5, the expression of endoplasmic reticulum stress proteins, including CCAAT/enhancer-binding protein homologous protein (CHOP), glucose-regulated protein 78 (GRP78), and cysteine-containing aspartate-specific proteases-12 (Caspase-12), and the expression of Wnt7/β-catenin in HBMECs. Results ·Compared with the Sham group, the brain water content of the infarction area of mice increased after MCAO, and the exudation of EB increased significantly (both P<0.05). The adhesion between cerebrovascular endothelial cells and pericytes in mice was reduced after the occurrence of MCAO. After intraperitoneal injection of 4-PBA in mice with MCAO, the degree of brain edema and the exudation of EB were reduced (both P<0.05), and the adhesion between cerebrovascular endothelial cells and pericytes increased. Compared with the HBMECs of the blank control group, the viability of HBMECs after OGD decreased, and the permeability of HBMECs increased (both P<0.05). OGD condition also led to decreased expression of connexins (ZO-1 and claudin-5), decreased secretion of PDGF-β in HBMECs, increased expression of endoplasmic reticulum stress proteins (CHOP, GRP78 and Caspase-12), up-regulated intracellular Ca2+ concentration and ROS content, and decreased expression of Wnt7 and β-catenin in HBMECs (all P<0.05). After HBMECs were cultured with 4-PBA, the damage of HBMECs caused by OGD was reduced, and the expression of connexins increased. The permeability of HBMECs was reduced, and the secretion of PDGF-β was promoted (all P<0.05). After 4-PBA treatment, the activity of Wnt7/β-catenin signaling was significantly restored in the OGD model of HBMECs (P<0.05). Conclusion ·Wnt7/β-catenin signaling inactivation caused by endothelial reticulum stress bursts leads to cerebrovascular endothelial cell damage, which is the crucial pathway of BBB destruction after stroke. Figures and Tables | References | Related Articles | Metrics

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Metformin ameliorates the mitochondrial damage induced by C9ORF72 amyotrophic lateral sclerosis/frontotemporal dementia-related poly-GR FENG Yiyuan, XU Zhongyun, YIN Yafu, WANG Hui, CHENG Weiwei 2023, 43 (7):  839-847.  doi: 10.3969/j.issn.1674-8115.2023.07.006 Abstract ( 243 )   HTML ( 19 )   PDF (4660KB) ( 258 )   Objective ·To investigate the effect of C9ORF72 amyotrophic lateral sclerosis/frontotemporal dementia (ALS/FTD)-related poly-glycine-arginine (poly-GR) on mitochondrial morphology and function, and analyze the rescue effect of metformin on mitochondrial damage induced by poly-GR and its underlying mechanism. Methods ·SK-N-SH cells stably overexpressing 50 repeated glycine-arginine sequences [(GR)50] or green fluorescent protein (GFP) were constructed by lentivirus infection, which were respectively named as (GR)50-SK cell line and GFP CTRL-SK cell line. (GR)50 expression in (GR)50-SK cells was verified by Western blotting. GFP expression in GFP GTRL-SK cells was observed by fluorescence microscope. Propidium iodide (PI) staining was used to detect the apoptosis levels of (GR)50-SK and GFP CTRL-SK cells. Immunofluorescence (IF) staining was performed to determine the subcellular location of (GR)50. Reactive oxygen species (ROS) level of mitochondria was evaluated by staining cells with MitoSOX Red followed by observing the intensity of red fluorescence under fluorescence microscope. The mitochondrial morphology of (GR)50-SK and GFP CTRL-SK cells was observed by transmission electron microscopy. Western blotting was used to detect protein kinase B (PKB, also known as AKT) and its phosphorylation levels in (GR)50-SK and GFP CTRL-SK cells. SC79 was used to activate AKT in (GR)50-SK cells, and MitoSOX Red staining and PI staining were used to analyze mitochondrial ROS and apoptosis levels after phosphorylated AKT increased. Metformin was used to treat (GR)50-SK and GFP CTRL-SK cells, respectively, and the apoptosis levels, mitochondrial ROS levels, mitochondrial morphology, AKT and its phosphorylation levels, and ATP concentrations of the two cells were detected by the above methods and ATP detection kit, respectively. Results ·Western blotting showed that the construction of (GR)50-SK cells was successful, and fluorescence microscopy showed that the construction of GFP CTRL-SK cells was also successful. PI staining results showed that the apoptosis level of (GR)50-SK cells was higher than that of the GFP CTRL-SK cells (P=0.016). IF staining results showed that there was partial co-localization of (GR)50 in the mitochondria of (GR)50-SK cells. Compared with GFP CTRL-SK cells, the mitochondrial morphology and structure of (GR)50-SK cells were significant abnormalities, with a significantly increased ROS levels. The AKT levels in (GR)50-SK cells were similar to those in the GFP CTRL-SK cells, but there was a significant decrease in phosphorylated AKT levels. After (GR)50-SK cells were treated with SC79, the AKT phosphorylation level was significantly upregulated, and ROS level and apoptosis level were significantly downregulated. Metformin could significantly up-regulate the phosphorylated AKT levels in (GR)50-SK cells, but had no effect on AKT levels; it could reshape the morphology and structure of some mitochondria, reduce ROS levels, increase ATP production (P=0.000), and down-regulate the level of cell apoptosis (P=0.000). Conclusion ·(GR)50 can cause mitochondrial morphology and function abnormalities by down-regulating AKT phosphorylation, and promote cell apoptosis. Metformin can effectively reduce the occurrence of the above pathological events induced by (GR)50. Figures and Tables | References | Related Articles | Metrics

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Establishment and phenotype verification of mouse oviductal epithelial organoids WU Qiqian, HU Yanqin, CHEN Jiale, LI Muchen, ZHAO Yougan, WU Jingwen 2023, 43 (7):  848-859.  doi: 10.3969/j.issn.1674-8115.2023.07.007 Abstract ( 374 )   HTML ( 21 )   PDF (8524KB) ( 286 )   Objective ·To establish a culture system of oviductal epithelial organoids from wild type (WT) mice and miR-34b/c-/- and miR-449-/- double knockout (dKO) mice, and verify the phenotypes. Methods ·The oviduct epithelial cells of WT mice and dKO mice were isolated and purified by enzyme digestion and differential adhesion method, and the purity of the isolated oviduct epithelial cells was identified by immunofluorescence staining. The numbers, growth rates and sizes of oviductal epithelial organoids between WT mice and dKO mice were compared by counting and diameter measurement. Hematoxylin-eosin (H-E) staining and transmission electron microscope (TEM) were used to observe the morphology and structure of the oviductal epithelial organoids. The proportions of ciliated cells and secretory cells in the oviductal epithelial organoids from WT mice and dKO mice were observed and counted by immunofluorescence staining. Immunohistochemistry (IHC), real-time quantitative PCR (RT-qPCR) and Western blotting were used to observe the expression levels of marker genes of ciliated cells and secretory cells in the oviductal epithelial organoids. Results ·The purity of the isolated and purified oviduct epithelial cells was high. Compared with the organoids from WT mice, the oviductal epithelial organoids from dKO mice grew faster and larger, and were more in number. But they developed more slowly than those from WT mice, as the invaginations of the dKO mice organoids appeared on the 28th day of culture, while the WT mice organoids exhibited the same structures on the 16th day. The oviductal epithelial organoids showed similar structures as those of the oviduct in vivo under hematoxylin-eosin (H-E) staining and TEM. Immunofluorescence staining showed that the ciliated cells of oviductal epithelial organoids from dKO mice were significantly reduced and the secretory cells were significantly increased (both P<0.05). IHC showed that the molecular expression patterns of the oviductal epithelial organoids were consistent with those of the oviducts in vivo, i.e. the expression levels of ciliated cell markers acetylated α-tubulin (Ac-α-tubulin) and forkhead box J1 (FOXJ1) decreased, and the expression level of the secretory cell marker paired box 8 (PAX8) increased. RT-qPCR showed that the mRNA levels of Foxj1 and tubulin β class Ⅳa (Tubb4a) decreased (both P<0.05), while Pax8 increased in the oviductal epithelial organoids of dKO mice (P<0.05). Western blotting results showed that the protein expression level of FOXJ1 in the organoids of dKO mice significantly decreased, while the expression of PAX8 significantly increased (both P<0.05). Conclusion ·The culture system of oviductal epithelial organoids of WT mice and dKO mice are successfully constructed, which can simulate the phenotypes of mouse oviduct in vivo. Figures and Tables | References | Related Articles | Metrics

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CXCL9 expression in breast cancer and its correlation with the characteristics of tumor immunoinfiltration DU Shaoqian, TAO Mengyu, CAO Yuan, WANG Hongxia, HU Xiaoqu, FAN Guangjian, ZANG Lijuan 2023, 43 (7):  860-872.  doi: 10.3969/j.issn.1674-8115.2023.07.008 Abstract ( 854 )   HTML ( 41 )   PDF (7698KB) ( 554 )   Objective ·To explore the effect of C-X-C motif chemokine ligand 9 (CXCL9) expression on the prognosis of breast cancer patients and its correlation with tumor-infiltrating immune cells (TIICs). Methods ·Transcriptome data of 1 100 breast tumor tissues and 112 adjacent tissues were obtained from The Cancer Genome Atlas (TCGA) database. CIBERSORT deconvolution algorithm was used to analyze the proportion of TIIC subgroups in breast cancer immune microenvironment and its effect on the prognosis of patients. Differentially expressed genes, immune-related genes and breast cancer prognostic-related genes were downloaded from TCGA database, ImmPort database and GEPIA2 data platform, respectively. The intersection relationships of the three gene sets were analyzed by using R language, and the target genes were screened. Based on the downloaded transcriptome data, CXCL9 positive-related genes,the difference of CXCL9 mRNA expression in breast cancer tissues and adjacent tissues and its effect on the prognosis of patients were analyzed. STRING data platform was used to analyze the protein-protein interaction (PPI) network of CXCL9. Gene Ontology (GO) function analysis and Kyoto Encyclopedia of Genes and Genome (KEGG) pathway analysis were performed on CXCL9 positive correlation genes and the genes corresponding to the interacting proteins obtained from the PPI network by using R language. Spearman correlation coefficient was used to analyze the correlation between CXCL9 mRNA expression and TIIC subgroups and immune checkpoint-related genes. Paraffin tissue samples of 60 clinical breast cancer patients were collected and made into tissue chips. The correlation between CXCL9 expression and CD8+ T cells infiltration in the tissue chips was detected by immunohistochemical staining (IHC). The types of CXCL9+ cells in breast cancer interstitium were analyzed by multiplex immunohistochemistry staining (mIHC). Kaplan-Meier (KM) survival curve was used to analyze the effect of CXCL9 mRNA expression and CD8+ T cell infiltration on the prognosis of breast cancer patients. Results ·CIBERSORT algorithm analysis showed that the distribution proportion of TIIC subgroups in breast cancer immune microenvironment varied greatly, and their effect on patients′ prognosis was also different. The Venn diagram of three types of gene sets was drawn, and CXCL9 was screened out. The top 150 positive correlation genes with CXCL9 were obtained. CXCL9 mRNA expression levels in four molecular types of breast cancer were higher than those in adjacent tissues (all P=0.000), and their high expressions were significantly associated with good prognosis of patients (P=0.013). A total of 41 interacting proteins were obtained through PPI network analysis. GO and KEGG analysis showed that CXCL9 and its related genes were mainly enriched in biological functions and pathways related to immune regulation. Spearman correlation coefficient analysis showed that the expression level of CXCL9 mRNA was positively correlated with CD8+ T cells infiltration ratio, negatively correlated with M2-type macrophages infiltration ratio, and positively correlated with most immune checkpoint genes expression (all P<0.05). IHC experiments showed that CXCL9 was highly expressed in breast cancer tissues compared with adjacent tissues, accompanied by an increased percentage of CD8+ T cells infiltration (P=0.000). mIHC results showed that CXCL9 was expressed in some CD68+ tumor-associated macrophages (TAMs) and CD11c+ dendritic cells (DCs) in the stroma of breast cancer. KM survival curve showed that when CXCL9 was highly expressed, CD8+ T cells high infiltration could prolong the survival of breast cancer patients. Conclusion ·CXCL9 can be used as a biomarker for good prognosis of breast cancer patients. The high expression of CXCL9 in the microenvironment of breast cancer is positively correlated with the infiltration ratio of CD8+ T cells and may activate its anti-tumor effect. The expression of CXCL9 may be closely related to the recruitment of lymphocytes into the tumor microenvironment for anti-tumor immune response. Figures and Tables | References | Related Articles | Metrics

Clinical research

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Role of 18F-MD-PSMA PET/CT in initial stage of intermediate and high risk prostate cancer YAN Yeqing, LIANG Sheng, YANG Bin, ZOU Renjian, MA Yufei, CAI Lisheng, WANG Hui, FU Hongliang 2023, 43 (7):  873-881.  doi: 10.3969/j.issn.1674-8115.2023.07.009 Abstract ( 302 )   HTML ( 16 )   PDF (2329KB) ( 160 )   Objective ·To evaluate the role of 18F-MD-PSMA PET/CT in the initial stage of patients with moderate and high risk prostate cancer (PCa). Methods ·A total of 67 patients with moderate and high risk PCa who were treated in Xinhua Hospital, Shanghai Jiao Tong University School of Medicine from September 2017 to June 2022 were initially staged by 18F-MD-PSMA PET/CT. Conventional imaging (CI), including multi-parameter magnetic resonance imaging (mp-MRI) and bone scintigraphy (BS), were performed within two weeks before 18F-MD-PSMA PET/CT. Twenty-five patients underwent 18F-FDG PET/CT at the same time. The sensitivity (SEN), specificity (SPEC), positive predictive value (PPV), negative predictive value (NPV) and accuracy (ACU) of 18F-MD-PSMA PET/CT in the initial stage were evaluated, and the results were compared with those of 18F-FDG PET/CT, mp-MRI and BS. The consistency of 18F-MD-PSMA PET/CT and CI in terms of primary lesion, regional lymph node metastasis and bone metastasis was evaluated by Kappa consistency test refering to the postoperative pathological T and N staging results and bone metastasis results of clinical follow-up. Kappa coefficient was calculated and compared. Results ·Of the 67 patients with PCa, 38 patients underwent radical prostatectomy and had completed pathological data, with 27 patients undergoing regional lymphadenectomy and 1 patient undergoing expanded pelvic lymphadenectomy at the same time. The pathological results were obtained as gold standard. The detection rates of mp-MRI and 18F-MD-PSMA PET/CT in diagnosing intrathecal lesions were both 100%. The SENs in diagnosing bilateral intralobular lesions were 26.3% and 63.2%, respctively; the SPECs were both 75.0%.The Kappa consistency test showed that the consistency of 18F-MD-PSMA PET/CT in diagnosis of extracapsular extension (EPE), seminal vesicle invasion (SVI), and bladder neck invasion (BNI) was higher than that of mp-MRI. Fisher′s exact test showed that there were no statistically significant differences in SEN (P=0.226, P=0.491) and SPEC (P=1.000, P=0.342) between the two methods for diagnosing EPE and SVI, as well as SEN (P=1.000) for diagnosing BNI. In terms of diagnosis of lymph node metastasis, based on the analysis of lymph node numbers, the consistency between 18F-MD-PSMA PET/CT and pathological results was higher than that of mp-MRI (Kappa coefficients of 0.555 and 0.137, respectively). Fisher′s exact test showed that there were no statistically significant differences in SEN and SPEC between the two examination methods (P=0.562, P=0.829). Based on the patients, the consistency between 18F-MD-PSMA PET/CT and pathological results was higher than that of mp-MRI (Kappa coefficients of 0.850 and 0.313, respectively). There was no statistically significant difference in SEN between the two methods (P=1.000). In terms of diagnosis of bone metastasis, based on the analysis of bone lesion numbers, the consistency between 18F-MD-PSMA PET/CT and clinical follow-up results was higher than that of BS (Kappa coefficients of 0.500 and 0.299, respectively). Fisher′s exact test showed that there was no statistically significant difference in SEN between the two methods (P=0.219). Based on the patients, the consistency between 18F-MD-PSMA PET/CT and clinical follow-up results was higher than that of BS (Kappa coefficients of 0.953 and 0.766, respectively). There was no statistically significant difference in SEN between the two methods (P=1.000). The risks of 21 patients (31.3%) were increased after 18F-MD-PSMA PET/CT detection, with 1 patient (1.5%) decreasing. The initial stage of 32 cases (47.8%) were changed after 18F-MD-PSMA PET/CT detection, with 27 cases (40.3%) upstaged and 5 cases (7.5%) downstaged. Conclusion ·18F-MD-PSMA PET/CT is superior to CI in the diagnosis of bilateral intralobular lesions, EPE, SVI, regional lymph node metastasis and bone metastasis in intermediate and high risk PCa, and on this basis, the diagnosis of clinical stage and metastatic status of some patients has been changed. Figures and Tables | References | Related Articles | Metrics

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Screening for pathogenic variants in obese cohort using whole-exome sequencing WANG Jinghui, ZHANG Hong, ZHANG Rong, PENG Danfeng, YU Hairong, CHEN Xianghui, XUAN Ye, HU Cheng, GU Yunjuan 2023, 43 (7):  882-889.  doi: 10.3969/j.issn.1674-8115.2023.07.010 Abstract ( 272 )   HTML ( 16 )   PDF (1911KB) ( 529 )   Objective ·To screen mutations of key genes in the leptin-melanocyte stimulating hormone (LEP-MSH) pathway by whole-exome sequencing (WES) in the obese cohort. Methods ·A total of 119 obese patients aged 17－65 years old with body mass index (BMI)≥34 kg/m2, who underwent laparoscopic sleeve gastrectomy from January 2011 to July 2019 at Shanghai Sixth People′s Hospital, Shanghai Jiao Tong University School of Medicine were collected. The peripheral blood samples of the research subjects were collected, and whole genome DNA was extracted to perform WES. Bioinformatic methods were applied to detect the mutations in 16 genes in the LEP-MSH pathway (ADCY3, AGRP, BDNF, KSR2, LEP, LEPR, MC3R, MC4R, MCHR1, MRAP2, NTRK2, PCSK1, PHIP, POMC, SH2B1,and SIM1). Rare variants with the minor allele frequency in the total population less than 0.02 and in the East Asian population less than 0.01 in the 1000 Genome (1000G), Exome Aggregation Consortium (ExAC) and Genome Aggregation Database (gnomAD) were selected for subsequent analysis. Six pieces of prediction software were used to evaluate the deleteriousness of the mutations. Finally, based on the clinical information of each patient, the pathogenicity of all variants was determined according to the guidelines of America College of Medical Genetics and Genomics (ACMG), and only the "pathogenic", "likely pathogenic", and "uncertain significance" variants were retained. Results ·A total of 26 variants, 22 kinds of variants were detected in 24 patients from 119 subjects, all of which were heterozygous mutations. The detected variants included 7 in SH2B1 gene (accounting for 26.92% of the total variants), 4 in MCHR1 gene (accounting for 15.38%), 3 in PHIP gene (accounting for 11.53%), 2 in ADCY3 and LEPR genes (accounting for 7.69%, respectively), and 1 in LEP, NTRK2, AGRP, KSR2, MC3R, MC4R, BDNF, and PCSK1 genes, respectively (accounting for 3.85%, respectively). There were 3 patients having the same mutation site in SH2B1 gene, and 2 patients having the same mutation sites in LEPR gene and MCHR1 gene, respectively. In addition, among these mutations, there were 12 ones not included in the East Asian population in 3 public databases, which were novel mutations in the East Asian population, located in SH2B1 (p.V209I, p.R67C, and p.L149F), KSR2 (p.P155T), LEP (p.D106N), LEPR (p.W132R), PHIP (p.K1461R), BDNF (p.N84S), PCSK1 (p.R282W), NTRK2 (p.T732M), MC3R (p.S71P), and MC4R (p.W174X). Conclusion ·A total of 22 kinds of rare variations possibly associated with obesity in the LEP-MSH pathway are detected, 12 of which are novel in the East Asian population. Figures and Tables | References | Related Articles | Metrics

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Study on long-term performance evaluation of auditory and speech ability in cochlear implant in congenital deaf children with cochlear nerve deficiency after cochlear implantation YANG Lu, HUANG Meiping, ZHOU Qian, LI Jin, LI Yun, HUANG Zhiwu 2023, 43 (7):  890-897.  doi: 10.3969/j.issn.1674-8115.2023.07.011 Abstract ( 261 )   HTML ( 20 )   PDF (1385KB) ( 188 )   Objective ·To evaluate the long-term performance and influencing factors of auditory and speech abilities, and social-life abilities in congenital deaf children with cochlear nerve deficiency (CND) after cochlear implant (CI) surgery. Methods ·Twenty-one CND children with CI implantation in Shanghai Ninth People′s Hospital, Shanghai Jiao Tong University School of Medicine were assigned to CND group, and twenty children with extremely severe sensorineural hearing loss with normal inner ear structure matching implantation age and CI use time of the CND group were selected as control group. The aided hearing threshold, Infant-toddler Meaningful Auditory Integration Scale (IT-MAIS), LittlEARS Auditory Questionnaire (LEAQ), Categories of Auditory Performance-Ⅱ (CAP-Ⅱ), Meaningful Use of Speech Scale (MUSS), Speech Intelligibility Rating (SIR), closed and open disyllabic speech recognition test, and speech intelligibility test were used to hierarchically evaluate the subjects′ auditory and verbal abilities, and the Infant-junior Middle School Student′s Social Life Ability Scale (S-M Scale) was used to assess the subjects′ social-life ability. Results ·The average aided hearing threshold of each frequency point in the CND group was significantly worse than that of the control group (all P<0.05). In the CND group, 90.5% of children′s aided hearing threshold of all frequencies entered the Chinese banana map. The auditory and speech abilities of the CND group were worse than those of the control group, and the significant differences between the two groups were found in the results of IT-MAIS, CAP-Ⅱ, MUSS, SIR, and closed and open disyllabic speech recognition (all P<0.05), but not in LEAQ. The social-life ability of the CND group was worse than that of the control group, and the difference was statistically significant (P<0.05). There was no correlation between implantation age, usage time of CI, auditory and language rehabilitation time, and auditory and speech ability of the CND group. Conclusion ·CI is an effective auditory intervention method for children with CND, but their average auditory and speech abilities and social-life abilities are significantly behind those of CI users with normal inner ear structure. Therefore, for CND children, individualized communication strategies and rehabilitation training methods should be emphasized, and the cultivation of social-life ability and psychological guidance should be strengthened. Figures and Tables | References | Related Articles | Metrics

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Identification of pathogenic mutations for a Wolfram syndrome pedigree by whole exome sequencing and analysis of its clinical characteristics MENG Xiangyu, YAN Dandan, CHEN Xianghui, LAI Siyu, XU Yun, GENG Ruina, ZHANG Hong, ZHANG Rong, HU Cheng, YAN Jing 2023, 43 (7):  898-905.  doi: 10.3969/j.issn.1674-8115.2023.07.012 Abstract ( 422 )   HTML ( 16 )   PDF (3177KB) ( 180 )   Objective ·To identify the causative gene and mutations and describe the clinical traits in a Chinese diabetes pedigree suspected of Wolfram syndrome. Methods ·A total of 12 subjects from one family were included. The proband was admitted to the Department of Endocrinology, The First Affiliated Hospital of Xinxiang Medical University, for the first time in May 2013. Then he visited the hospital for follow-up in July 2022 and in April 2023, respectively. The other members of this family included the proband′s sister, father, mother, paternal grandfather, paternal grandmother, uncle, aunt, as well as maternal grandfather, maternal grandmother, and two brothers of the proband′s mother. Clinical data of all subjects were collected. The whole exome sequencing was used to screen the pathogenic genes and mutation sites of six members of the family, and Sanger sequencing was used to verify the above results. Effects of the mutation of the pathogenic gene WFS1 in Wolfram syndrome on the function of the wolframin protein were evaluated by bioinformatics softwares, including CADD, DANN, MetaSVM, Polyphen-2, SIFT and M-CAP. The three-dimensional structures of wild-type and mutant wolframin proteins were constructed with Swiss-Model software, and visualized with PyMOL software. Cluster Omega software was used for evaluating species conservation of WFS1 gene mutation sites. JNetPRED software was used for online prediction of wolframin protein secondary structure. Results ·The proband and his sister both carried R558H and S411Cfs*131 mutations, two compound heterozygous mutations of the Wolfram syndrome pathogenic gene WFS1. The proband′s father and parental grandfather both carried the R558H mutation, while the proband′s mother and maternal grandfather both carried the S411Cfs*131 mutation. The R558H mutation was a rare missense mutation, and the S411Cfs*131 mutation was a novel frameshift mutation. Bioinformatics analysis softwares predicted that the R558H mutation located in the α-helical structure of the wolframin protein. This mutation was a damage mutation and the amino acid sequence of the mutation region was highly conservative among 12 species with varying degrees of evolution, including humans. Conclusion ·Two causative mutations of WFS1 gene are identified in a Chinese diabetes pedigree by whole exome sequencing. The study supplements the existing genotype and phenotype profiles of Wolfram syndrome, which can realize early diagnosis of diabetes pedigrees and help in performing timely follow-up of patients, so as to achieve early intervention and treatment of this disease. Figures and Tables | References | Related Articles | Metrics

Evidence-based medicine

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A meta-analysis of the effects of levothyroxine dose adjustment on maternal and infant outcomes in pregnant women with hypothyroidism CHEN Hui, ZHU Weiyi, YAO Yijin 2023, 43 (7):  906-915.  doi: 10.3969/j.issn.1674-8115.2023.07.013 Abstract ( 189 )   HTML ( 22 )   PDF (4423KB) ( 102 )   Objective ·To evaluate the effects of levothyroxine (L-T4) dose adjustment according to the level of thyroid stimulating hormone (TSH) on maternal and infant outcomes in the pregnant women with hypothyroidism by meta-analysis. Methods ·China National Knowledge Infrastructure (CNKI), China Science and Technology Journal Database (VIP), Wanfang Data Knowledge Service Platform, PubMed, Cochrane Library and Embase were retrieved to collect all the controlled studies on the treatment of pregnant women with hypothyroidism by adjusting the dose of L-T4 according to TSH level from the establishment of the databases to April 9, 2022. The references were also traced. Literature screening, data extraction, and quality evaluation were performed independently by two researchers. Cochrane evaluation was used to evaluate the quality of the included literature. Outcome indicators included gestational hypertension, gestational diabetes, postpartum hemorrhage, delivery mode, preterm birth, fetal death, neonatal asphyxia, and low birth weight infants. RevMan 5.3 was used for meta-analysis. Result ·A total of 1 268 articles were retrieved from 6 databases, and 8 were included in the study, including 4 Chinese articles and 4 English articles. The overall risk of study bias was at a moderate level. Compared with the control group, the OR of gestational diabetes risk was 0.61 (95%CI 0.44?0.86, P=0.004) and the OR of fetal death risk was 0.38 (95%CI 0.18?0.81, P=0.010) in the experimental group with L-T4 dose adjusted according to the TSH level of the pregnant women with hypothyroidism, which were both statistically significant. However, the treatment method of adjusting L-T4 dose did not affect the risks of vaginal delivery [OR=1.82 (95%CI 0.75?4.40, P=0.180)], gestational hypertension [OR=0.77 (95%CI 0.53?1.12, P=0.170)], postpartum hemorrhage [OR=1.20 (95%CI 0.50?2.92, P=0.680)], preterm birth [OR=0.72 (95%CI 0.48?1.06, P=0.100)], low birth weight infants [OR=1.00 (95%CI 0.65?1.54, P=0.999)], or neonatal asphyxia [OR=0.50 (95%CI 0.20?1.27, P=0.150)] significantly. Conclusion ·Adjusting the L-T4 therapeutic dose according to the TSH level may help reduce the risks of gestational diabetes and fetal death in the pregnant women with hypothyroidism. Figures and Tables | References | Related Articles | Metrics

Review

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Research progress of exercise therapy for depressive disorder ZHANG Shuoyuan, LI Chunbo 2023, 43 (7):  916-922.  doi: 10.3969/j.issn.1674-8115.2023.07.014 Abstract ( 490 )   HTML ( 25 )   PDF (1341KB) ( 468 )   Depressive disorder, as one of the major diseases in the world, has always received much attention for its prevention and treatment. As an emerging treatment, exercise therapy has optimistic application prospect with the advantages such as lower cost, fewer side effects and easier implementation, compared to conventional treatments such as drug therapy and physical therapy. Relevant studies have explored the mechanism of exercise in the treatment of depressive disorder, but the mechanism is not clear yet, which may involve improving the levels of neurobiochemical molecules, inhibiting inflammatory response, regulating neuroendocrine system, improving neuroplasticity, and other aspects. Exercise therapy has been proved to have similar biological effects with antidepressants, and may have overlapping effects with other treatments. Early intervention can benefit both non- diseased and already diseased populations to a certain extent. At present, there is still a gap in the clinical field related to exercise therapy for depressive disorder, and there are few high-quality studies. The design of exercise therapy plans is still in the exploratory stage, and there is no consensus on the design of exercise therapy plans. Additionally, there is a lack of relevant exercise therapy guidelines for clinicians to refer to. This review systematically introduces the biological mechanism of exercise therapy for depressive disorder, summarizes the clinical research results in this field carried out at home and abroad, and analyzes the current program and advantages and disadvantages of exercise therapy, in order to provide reference for the in-depth development of the exercise therapy researches. References | Related Articles | Metrics

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Application progress of CT radiomics in gastrointestinal stromal tumor MA Ben, ZHAO Cheng, SHU Yijun, DONG Ping 2023, 43 (7):  923-930.  doi: 10.3969/j.issn.1674-8115.2023.07.015 Abstract ( 280 )   HTML ( 12 )   PDF (1786KB) ( 249 )   Gastrointestinal stromal tumor (GIST) is the most common mesenchymal tumor in the gastrointestinal tract, with complex biological characteristics and varying risks, and the treatment methods and prognosis of patients with different risks are quite different; therefore, early diagnosis and risk assessment are crucial for its precision treatment. In recent years, CT radiomics, as an emerging imaging technology, can transform traditional CT image features into a large number of data, thereby reflecting the inherent heterogeneity of GIST and even correlating with its gene expression features. This paper reviews the research progress of CT radiomics in the diagnosis and prediction of GIST with the help of machine learning. The current CT radiomics can not only be used for the differential diagnosis of GIST and other gastric diseases, but also for the risk evaluation of GIST. Furthermore, pathological analysis and gene diagnosis can be performed based on CT images, and then the first-line treatment effect and long-term prognosis can be predicted. At present, various prediction models constructed by combination of CT radiomics and clinical information have been well verified in the specific practice of different clinical problems, showing broad application prospects. However, in the specific clinical application process, different methods of sample data collection and processing, differences in the selection of machine learning algorithms, and the selection of 2D or 3D images all affect the specific effectiveness of CT radiomics. Hence, unified and standardized application rules for radiomics has to be established. Figures and Tables | References | Related Articles | Metrics

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Research progress of immune response regulated by epigenetic modification in pneumonia WANG Qing, HAN Xiao, ZHANG Xiaobo 2023, 43 (7):  931-938.  doi: 10.3969/j.issn.1674-8115.2023.07.016 Abstract ( 285 )   HTML ( 21 )   PDF (1688KB) ( 462 )   Pneumonia is one of the most common infectious diseases, and although considerable progress has been made in the diagnosis and treatment of pneumonia, it is still associated with high mortality, prolonged hospitalization, and significant medical expenditures. Epigenetic modifications are heritable changes in gene expression without altering the DNA sequence, including DNA methylation, histone modification, non-coding RNA and RNA modification, which are involved in regulating gene expression at multiple levels, including DNA, histone, and transcriptional and post-transcriptional levels. A growing number of studies have suggested that epigenetic regulation may play a central role in the initiation and progression of pneumonia by regulating the immune function. Following the infection with pathogens in the lungs, epigenetic modification can affect the occurrence and progression of pneumonia in different individuals by regulating the inflammatory and immune response, including the development and differentiation of various immune cells, the recognition and transduction of infection signals, and the production of cytokines and anti-pathogen effector molecules. By summarizing recent studies on epigenetic modification of immunity in pneumonia, this review elucidates the key role that epigenetic modification of immunology plays in the initiation and progression of pneumonia, as well as its potential application to clinical diagnosis and therapeutic targets in the treatment of pneumonia, providing a sound theoretical basis for further research. Figures and Tables | References | Related Articles | Metrics

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Research progress of brain magnetic resonance imaging related to suicide in bipolar disorder patients CHEN Jiaye, ZHANG Huifeng, LU Wenxian, PENG Daihui 2023, 43 (7):  939-944.  doi: 10.3969/j.issn.1674-8115.2023.07.017 Abstract ( 236 )   HTML ( 20 )   PDF (1288KB) ( 207 )   Bipolar disorder, as a major mental illness, has a high lifetime suicide attempt rate in patients, and suicidal behavior is most likely to occur during depressive episodes. Therefore, in-depth study of its mechanism is essential for prevention, early detection and intervention of suicide. With the development of magnetic resonance imaging (MRI) technology, it has been found that there are abnormalities in the brain structure and function in suicidal patients with bipolar disorder. This article reviews the studies on suicide in bipolar disorder patients by MRI from four aspects: structure, function, structure-function, and central metabolism and cerebral blood flow perfusion, and summarizes the suicide-related changes. This review focuses on distinguishing the brain MRI changes under different mood states and diverse definitions of suicide, aiming to provide reference for further exploration of the pathophysiological mechanism of suicide in bipolar disorder. References | Related Articles | Metrics