Objective:To optimize the laboratory methods for isolation, culture, and preservation of Neisseria gonorrhoeae ( N. gonorrhoeae) . Methods:Five quality control bacterial strains ( N. gonorrhoeae strain 1, N. gonorrhoeae strain 2, Neisseria mucosa strain 3, Enterococcus faecalis strain 4, and mixed bacterial strain 5) were separately cultured using a self-made gonococcal selective medium and 7 commercialized gonococcal selective media (gonococcal isolation plates, gonococcal T-M selective medium, gonococcal chocolate agar plates, gonococcal selective medium, modified Thayer-Martin [MTM] agar plates, disposable gonococcus [GC] agar medium, and gonococcal agar plates), and their cultivation performance was evaluated. Sixteen strains of N. gonorrhoeae were cultured both inside and outside a candle jar to screen for CO 2-dependent strains. The preservation performance of 4 self-made gonococcal preservation solutions, including calf serum with 10% dimethyl sulfoxide, calf serum with 10% glycerol, brain-heart infusion broth with 20% glycerol, and trypticase soy broth with 30% glycerol, was evaluated. The survival of N. gonorrhoeae in freeze-dried and non-freeze-dried states was observed. Results:The growth performance of the 5 quality control strains varied across different commercialized gonococcal culture media. Concretely, N. gonorrhoeae strain 1 formed large and numerous colonies on both the self-made culture medium and MTM agar plates, which outperformed the other 6 culture media; the growth performance of N. gonorrhoeae strain 2 on the 7 commercialized culture media was inferior to that on the self-made culture medium; all 7 commercialized culture media had inhibitory effects on the growth of Neisseria mucosa strain 3, among which the self-made culture medium, gonococcal chocolate agar plates, MTM agar plates, disposable GC agar medium, and gonococcal agar plates could completely inhibit its growth; the gonococcal T-M selective medium could completely inhibit the growth of Enterococcus faecalis strain 4; mixed bacterial strain 5 showed better separation performance on the self-made culture medium, gonococcal isolation plates, gonococcal T-M selective medium, gonococcal chocolate agar plates, MTM agar plates, and disposable GC agar medium. Under CO 2-enriched conditions, all 16 strains of N. gonorrhoeae exhibited good growth performance; however, the growth of 11 strains was markedly inhibited without CO 2. No significant differences were observed in the preservation performance of the 4 preservation solutions at -70°C, and confluent colonies could be observed in all preservation solutions after 12 months of strain preservation; at -20 ℃, the trypticase soy broth with 30% glycerol showed the best preservation performance, with a few viable strains remaining after 6 months, while the calf serum with 10% dimethyl sulfoxide performed worst, with partial strains remaining viable after 2 weeks. Non-freeze-dried N. gonorrhoeae survived for varying duration at different temperatures (4 ℃, -18 ℃, -29 ℃, -70 ℃, and liquid nitrogen) ; no N. gonorrhoeae strains survived by day 4 when stored at 4°C; freeze-dried N. gonorrhoeae remained viable with the presence of confluent colonies for 6 months at 4 ℃. Conclusion:The self-made gonococcal selective medium demonstrated superior cultivation and isolation performance compared to commercialized gonococcal selective media; a small amount of CO 2 could promote the growth of N. gonorrhoeae; ultralow temperature and freeze-drying preservation could increase the survival time of N. gonorrhoeae, with freeze-drying at 4 ℃ being the most cost-effective long-term preservation method.

BACKGROUND:The pathogenesis of rheumatoid arthritis has not yet been fully clarified,and recent studies have shown that mitophagy is associated with rheumatoid arthritis,but the key mechanisms need to be explored in depth.OBJECTIVE:To identify and validate the core interaction genes of mitophagy in rheumatoid arthritis using multiple machine learning algorithms and to analyze its immunoregulatory process.METHODS:The rheumatoid arthritis transcriptome expression dataset GSE15573 was retrieved from the GEO database as an independent training set,with the GSE97779 and GSE55235 collections used as independent validation sets.The differentially expressed genes of rheumatoid arthritis were screened using the training set,and"WGCNA"analysis was also performed.Then we downloaded the mitophagy-related genes from the"MitoCarta3.0"database,and intersected them with the differentially expressed genes of rheumatoid arthritis and the genes in the"WGCNA"analysis module to obtain the rheumatoid arthritis-mitophagy-related genes,and then analyzed the related genes for functional enrichment to clarify the cellular pathways.Feature genes were initially identified using the"Random Forest"and"Lasso"algorithms.The overlapping genes from these two methods were further refined using the"GMM"algorithm to identify the core interaction genes between rheumatoid arthritis and mitophagy.A predictive model was then developed and validated using an external dataset.Finally,"CIBERSORT"was employed to analyze the proportions and interactions of immune cell subsets during immune infiltration,while"ssGSEA"was used to examine the associations between the rheumatoid arthritis-mitophagy core interaction genes and immune cell subsets."ssGSEA"was also utilized to analyze the"GO"and"KEGG"biological pathways of the core interaction genes.RESULTS AND CONCLUSION:(1)Totally 807 differentially expressed genes in rheumatoid arthritis were obtained by differential analysis,1 208 genes were selected from two feature modules by"WGCNA"analysis,1136 genes were sorted out from the MitoCarta 3.0 database,and 53 HUB genes were obtained from the intersection of the three genes as rheumatoid arthritis-mitophagy related genes.(2)The results of functional enrichment analysis of related genes showed that the cellular processes were mainly related to mitophagy,peroxisome metabolism,cellular senescence,and necroptosis.(3)The three machine learning algorithms identified four rheumatoid arthritis-mitophagy core interaction genes(DNAJA3,C12orf65,AKR7A2,and PDHB).The area under the curve of nomoscore was 0.989,and the area under the curve values of rheumatoid arthritis-mitophagy core interaction genes verified by the receiver operating characteristic curve of external patient samples were all greater than 0.7.(5)Immunoregulatory analysis showed that the mitophagy process in rheumatoid arthritis was closely associated with memory B cells,M0 macrophages,activated memory CD4 T cells,and resting memory CD4 T cells.(6)The biological pathway analysis revealed that the core interaction genes were strongly associated with 821"GO"pathways(|cor|＞0.8,P＜0.001)and 48"KEGG"pathways(|cor|＞0.8,P＜0.001).The key biological processes identified were related to mitochondrial DNA metabolic process,mitochondrial DNA repair,mitochondrial DNA replication,mitochondrial genome maintenance,positive regulation of mitochondrial depolarization,and positive regulation of mitochondrial outer membrane permeabilization involved in apoptotic signaling pathway.To conclude,DNAJA3,C12orf65,AKR7A2,and PDHB are the core interaction genes of the mitophagy process in rheumatoid arthritis,which play key roles in disease progression by participating in specific immune processes and have precise and predictive effects on the diagnosis of rheumatoid arthritis.

BACKGROUND:Mitochondrial autophagy is closely related to the occurrence and development of steroid-induced osteonecrosis of the femoral head(SONFH),but specific biomarkers and regulatory mechanisms remain unclear. OBJECTIVE:To identify the key biomarkers of mitochondrial autophagy in steroid-induced osteonecrosis of the femoral head using machine learning algorithms and to conduct an immune infiltration analysis. METHODS:The SONFH datasets GSE123568 and GSE74089 were downloaded from the GEO database,serving as the training and validation sets,respectively.Differentially expressed genes between SONFH and control groups were selected,and weighted gene co-expression network analysis was performed.Mitochondrial autophagy-related genes were obtained from MitoCarta3.0 and intersected with differentially expressed genes and module genes.Two machine learning algorithms were utilized to identify key genes of SONFH mitochondrial autophagy,and validated using an external validation set.CIBERSORT and immune infiltration analysis were employed to assess the proportion of immune cells,and ssGSEA was used to analyze the correlation between mitochondrial autophagy genes and immune cells. RESULTS AND CONCLUSION:Differential analysis identified a total of 1 163 differentially expressed genes,including 663 upregulated genes and 500 downregulated genes.Weighted gene co-expression network analysis identified 4 key modules,comprising 1 412 module genes.Intersection with mitochondrial autophagy genes yielded 39 intersecting genes as disease-related mitochondrial autophagy genes.Gene ontology enrichment analysis showed that the biological processes were mainly related to heme metabolism,mitochondrial transport,nucleotide bisphosphate metabolism and thioester metabolism,and the cellular components were mainly related to mitochondrial matrix,mitochondrial outer membrane,organelle outer membrane and mitochondrial inner membrane,and the molecular functions were mainly related to fatty acid ligase activity,iron-sulfur cluster binding,and cofactor A ligase activity.Kyoto Encyclopedia of Genes and Genomes enrichment analysis mapped out a total of six pathways,which were mainly related to fatty acid degradation,mitochondrial autophagy,butyric acid metabolism,fatty acid biosynthesis and cofactor biosynthesis.Through LASSO regression and RFE-SVM algorithm analysis,four intersecting genes(ALDH5A1,FBXL4,MCL1,and STOM)were identified.The receiver operating characteristic curves of the four core genes and the diagnostic column chart validation set were all greater than 0.9.The occurrence and development of SONFH were related to immune cells such as dendritic cells,bone marrow-derived suppressor cells,regulatory T cells,and central memory CD8 T cells.To conclude,the four key mitochondrial autophagy genes ALDH5A1,FBXL4,MCL1,and STOM play a crucial role in the progression of SONFH through osteoclast differentiation and immune mechanisms.Additionally,all four genes have good disease prediction efficacy and can serve as biomarkers for the diagnosis and treatment of SONFH.

BACKGROUND:Our previous research found that Panax notoginseng can repair the morphological structure of bone cells,which has a good application prospect in the treatment of osteoarthritis,but the specific mechanism of Panax notoginseng is still unclear. OBJECTIVE:To identify the main components of Panax notoginseng using ultra-high performance liquid chromatography-Q exactive-mass spectrometry(UHPLC-QE-MS),and to explore the mechanism of Panax notoginseng in the treatment of osteoarthritis by combining network pharmacology,molecular docking and molecular dynamics simulation. METHODS:After identifying the main components of Panax notoginseng by UHPLC-QE-MS technology,the active components were screened by TCMSP database,and the targets of active components were found by TCMSP and Uniprot database.Osteoarthritis targets were screened out through disease databases.After the intersection of drug targets and disease targets,the protein-protein interaction network was constructed by importing STRING database and Cytoscape software,and the"active ingredient-action target"network was constructed to screen key active ingredients.Then the key targets were enriched and analyzed,and the key active components and key targets were verified by molecular docking.Finally,the results with the lowest binding energy were selected for molecular dynamics simulation. RESULTS AND CONCLUSION:A total of 57 active components were identified in the solution of Panax Notoginseng,including 50 intersection targets of components and disease targets,5 key active components(quercetin,ursodeoxycholic acid,kaempferol,naringenin and erythrocyanine),and 5 key targets(interleukin 6,matrix metalloproteinase 9,interleukin 1β,albumin and recombinant chemokine c-motif ligand 2).Gene ontology enriched 642 entries,among which 620 entries represent biological processes,21 entries represent molecular functions,and 1 entry represents cellular components.Kyoto encyclopedia of genes and genomes analysis indicated 63 pathways,mainly including estrogen signaling pathway,interleukin 17 signaling pathway and hyperglycosylation end product-hyperglycosylation end product receptor signaling pathway.Molecular docking showed good binding activity of key active components and key targets.Molecular dynamics simulation indicated that the stable interaction between quercetin and matrix metalloproteinase 9.The composition of Panax notoginseng was comprehensively studied,and the material basis of its efficacy was preliminarily clarified.It was predicted that Panax notoginseng could play an anti-inflammatory,cartilage-protective,and immunomodulatory role in treating osteoarthritis through multiple components,targets,approaches and pathways.

Osteoporosis （OP） is a metabolic disorder characterized by microstructural deterioration of bone and increased bone fragility due to reduced bone mass， which can cause the development of bone-related diseases. This condition imposes significant economic and psychological burdens on patients. While modern medicine has extensively researched the pathogenesis of OP， it remains incompletely understood. Current clinical management primarily relies on anti-resorptive drugs and synthetic metabolic agents. However， long-term use of some medications may yield suboptimal therapeutic outcomes and lead to severe adverse reactions. Given the necessity for prolonged or lifelong treatment for OP， there is a critical need to identify highly effective， safe， and cost-effective pharmaceutical interventions. In light of evolving disease management paradigms and recent advancements in OP research， traditional Chinese medicine has demonstrated emerging advantages in addressing this condition. Through literature review， this study delves into the pathogenesis of OP from five perspectives： hormonal dysregulation， autophagy， ferroptosis， oxidative stress， and intestinal flora alteration. Furthermore， it summarizes the therapeutic efficacy and specific mechanisms of traditional Chinese medicine monomers and compound formulas against OP through regulating hormone levels， interfering with autophagy， inhibiting ferroptosis， counteract oxidative stress，and maintain intestinal flora balance. These multifaceted insights are expected to provide theoretical reference and guide future clinical traditional Chinese medicine approaches for preventing and managing OP.

Objective:To investigate the long-term efficacy, safety and prognostic factors of intraoperative radiotherapy (IORT) for narrow-margin (resection margin < 1 cm) hepatectomy in patients with hepatocellular carcinoma (HCC) during radical surgery.Methods:A retrospective cohort study was conducted. The data of primary HCC patients undergoing radical surgery and narrow-margin hepatectomy IORT in the Cancer Hospital of the Chinese Academy of Medical Sciences from November 2009 to February 2019 were collected. IORT applied 6 MeV or 9 MeV electron beams and a single irradiation was given to the margin. Kaplan-Meier method was used for the overall survival (OS) and disease-free survival (DFS) analysis; log-rank test was used for survival comparison among subgroups. The recurrence patterns and adverse reactions were recorded. Univariate and multivariate Cox proportional hazards models were used to analyze the factors influencing the OS and DFS.Results:A total of 64 patients were enrolled, with the median age [ M ( Q1, Q3)] of 57 years (49, 63) years. All patients included 55 males (85.9%) and 9 females (14.1%). The median dose of IORT was 15 Gy (range: 12-17 Gy). The median follow-up time was 83.3 (64.4, 91.9) months. The 1-year, 3-year, 5-year, 7-year, 10-year OS rates were 90.4%, 80.6%, 75.5%, 71.4% and 47.6%, respectively; the 1-year, 3-year, 5-year, 7-year,10-year DFS rates were 77.8%, 68.1%, 59.6%, 57.6% and 38.4%, respectively. Univariate Cox regression analysis indicated that preoperative serum alpha-fetoprotein (AFP) > 400 ng/ml was an independent risk factor for poor OS (> 400 ng/ml vs. ≤ 400 ng/ml: HR = 6.57, 95% CI: 2.16-19.96, P < 0.001), while not the independent influencing factor of poor DFS ( HR = 1.71, 95% CI: 0.65-4.52, P = 0.277). The age ≤ 60 years or not, gender, viral hepatitis or not, American Joint Committee on Cancer stage, tumor diameter (> 5 cm or not), tumor number, degree of tumor differentiation, microvascular invasion or not, microsatellite nodules or not, anatomical liver resection or not, and the dose of IORT ≤15 Gy or not were not the independent influencing factors of poor OS and DFS (all P > 0.05). Kaplan-Meier method analysis showed that patients with preoperative serum AFP ≤ 400 ng/ml (48 cases) had better OS compared with those with preoperative serum AFP>400 ng/ml (16 cases) (5-year OS rate: 84.8% vs. 44.9%; 7-year OS rate: 79.9% vs.37.4%), and the difference was statistically significant ( P = 0.002). There was no statistically significant difference in the DFS between the 2 groups ( P = 0.134). During the follow-up, 28 patients (43.8%) relapsed, including 17 cases (26.6%) of early recurrence and 11 cases (17.2%) of late recurrence. No marginal recurrence was observed. There were 22 cases (34.4%) of intrahepatic recurrence alone, 2 cases (3.1%) of extrahepatic recurrence and 4 cases (6.3%) of stimutaneous recurrence inside and outside the liver. The 1-, 3-, 5- and 7-year cumulative recurrence rates inside the liver were 19.0%, 27.2%, 37.4% and 39.3% respectively, and the cumulative recurrence rates outside the liver were 6.4%, 8.0%, 9.6% and 9.6% respectively. There were no adverse reactions above grade 3 in the entire group. There were no surgery-related deaths within 30 d after the operation, and no radiation-induced liver disease occurred. Conclusions:Narrow-margin IORT helps HCC patients receiving hepatectomy to achieve favorable long-term survival and adverse reactions are tolerable. It can be used as a safe and effective adjuvant therapy alternative.

Objective:To optimize the laboratory methods for isolation, culture, and preservation of Neisseria gonorrhoeae ( N. gonorrhoeae) . Methods:Five quality control bacterial strains ( N. gonorrhoeae strain 1, N. gonorrhoeae strain 2, Neisseria mucosa strain 3, Enterococcus faecalis strain 4, and mixed bacterial strain 5) were separately cultured using a self-made gonococcal selective medium and 7 commercialized gonococcal selective media (gonococcal isolation plates, gonococcal T-M selective medium, gonococcal chocolate agar plates, gonococcal selective medium, modified Thayer-Martin [MTM] agar plates, disposable gonococcus [GC] agar medium, and gonococcal agar plates), and their cultivation performance was evaluated. Sixteen strains of N. gonorrhoeae were cultured both inside and outside a candle jar to screen for CO 2-dependent strains. The preservation performance of 4 self-made gonococcal preservation solutions, including calf serum with 10% dimethyl sulfoxide, calf serum with 10% glycerol, brain-heart infusion broth with 20% glycerol, and trypticase soy broth with 30% glycerol, was evaluated. The survival of N. gonorrhoeae in freeze-dried and non-freeze-dried states was observed. Results:The growth performance of the 5 quality control strains varied across different commercialized gonococcal culture media. Concretely, N. gonorrhoeae strain 1 formed large and numerous colonies on both the self-made culture medium and MTM agar plates, which outperformed the other 6 culture media; the growth performance of N. gonorrhoeae strain 2 on the 7 commercialized culture media was inferior to that on the self-made culture medium; all 7 commercialized culture media had inhibitory effects on the growth of Neisseria mucosa strain 3, among which the self-made culture medium, gonococcal chocolate agar plates, MTM agar plates, disposable GC agar medium, and gonococcal agar plates could completely inhibit its growth; the gonococcal T-M selective medium could completely inhibit the growth of Enterococcus faecalis strain 4; mixed bacterial strain 5 showed better separation performance on the self-made culture medium, gonococcal isolation plates, gonococcal T-M selective medium, gonococcal chocolate agar plates, MTM agar plates, and disposable GC agar medium. Under CO 2-enriched conditions, all 16 strains of N. gonorrhoeae exhibited good growth performance; however, the growth of 11 strains was markedly inhibited without CO 2. No significant differences were observed in the preservation performance of the 4 preservation solutions at -70°C, and confluent colonies could be observed in all preservation solutions after 12 months of strain preservation; at -20 ℃, the trypticase soy broth with 30% glycerol showed the best preservation performance, with a few viable strains remaining after 6 months, while the calf serum with 10% dimethyl sulfoxide performed worst, with partial strains remaining viable after 2 weeks. Non-freeze-dried N. gonorrhoeae survived for varying duration at different temperatures (4 ℃, -18 ℃, -29 ℃, -70 ℃, and liquid nitrogen) ; no N. gonorrhoeae strains survived by day 4 when stored at 4°C; freeze-dried N. gonorrhoeae remained viable with the presence of confluent colonies for 6 months at 4 ℃. Conclusion:The self-made gonococcal selective medium demonstrated superior cultivation and isolation performance compared to commercialized gonococcal selective media; a small amount of CO 2 could promote the growth of N. gonorrhoeae; ultralow temperature and freeze-drying preservation could increase the survival time of N. gonorrhoeae, with freeze-drying at 4 ℃ being the most cost-effective long-term preservation method.

BACKGROUND:The pathogenesis of rheumatoid arthritis has not yet been fully clarified,and recent studies have shown that mitophagy is associated with rheumatoid arthritis,but the key mechanisms need to be explored in depth.OBJECTIVE:To identify and validate the core interaction genes of mitophagy in rheumatoid arthritis using multiple machine learning algorithms and to analyze its immunoregulatory process.METHODS:The rheumatoid arthritis transcriptome expression dataset GSE15573 was retrieved from the GEO database as an independent training set,with the GSE97779 and GSE55235 collections used as independent validation sets.The differentially expressed genes of rheumatoid arthritis were screened using the training set,and"WGCNA"analysis was also performed.Then we downloaded the mitophagy-related genes from the"MitoCarta3.0"database,and intersected them with the differentially expressed genes of rheumatoid arthritis and the genes in the"WGCNA"analysis module to obtain the rheumatoid arthritis-mitophagy-related genes,and then analyzed the related genes for functional enrichment to clarify the cellular pathways.Feature genes were initially identified using the"Random Forest"and"Lasso"algorithms.The overlapping genes from these two methods were further refined using the"GMM"algorithm to identify the core interaction genes between rheumatoid arthritis and mitophagy.A predictive model was then developed and validated using an external dataset.Finally,"CIBERSORT"was employed to analyze the proportions and interactions of immune cell subsets during immune infiltration,while"ssGSEA"was used to examine the associations between the rheumatoid arthritis-mitophagy core interaction genes and immune cell subsets."ssGSEA"was also utilized to analyze the"GO"and"KEGG"biological pathways of the core interaction genes.RESULTS AND CONCLUSION:(1)Totally 807 differentially expressed genes in rheumatoid arthritis were obtained by differential analysis,1 208 genes were selected from two feature modules by"WGCNA"analysis,1136 genes were sorted out from the MitoCarta 3.0 database,and 53 HUB genes were obtained from the intersection of the three genes as rheumatoid arthritis-mitophagy related genes.(2)The results of functional enrichment analysis of related genes showed that the cellular processes were mainly related to mitophagy,peroxisome metabolism,cellular senescence,and necroptosis.(3)The three machine learning algorithms identified four rheumatoid arthritis-mitophagy core interaction genes(DNAJA3,C12orf65,AKR7A2,and PDHB).The area under the curve of nomoscore was 0.989,and the area under the curve values of rheumatoid arthritis-mitophagy core interaction genes verified by the receiver operating characteristic curve of external patient samples were all greater than 0.7.(5)Immunoregulatory analysis showed that the mitophagy process in rheumatoid arthritis was closely associated with memory B cells,M0 macrophages,activated memory CD4 T cells,and resting memory CD4 T cells.(6)The biological pathway analysis revealed that the core interaction genes were strongly associated with 821"GO"pathways(|cor|＞0.8,P＜0.001)and 48"KEGG"pathways(|cor|＞0.8,P＜0.001).The key biological processes identified were related to mitochondrial DNA metabolic process,mitochondrial DNA repair,mitochondrial DNA replication,mitochondrial genome maintenance,positive regulation of mitochondrial depolarization,and positive regulation of mitochondrial outer membrane permeabilization involved in apoptotic signaling pathway.To conclude,DNAJA3,C12orf65,AKR7A2,and PDHB are the core interaction genes of the mitophagy process in rheumatoid arthritis,which play key roles in disease progression by participating in specific immune processes and have precise and predictive effects on the diagnosis of rheumatoid arthritis.

Objective:To investigate the long-term efficacy, safety and prognostic factors of intraoperative radiotherapy (IORT) for narrow-margin (resection margin < 1 cm) hepatectomy in patients with hepatocellular carcinoma (HCC) during radical surgery.Methods:A retrospective cohort study was conducted. The data of primary HCC patients undergoing radical surgery and narrow-margin hepatectomy IORT in the Cancer Hospital of the Chinese Academy of Medical Sciences from November 2009 to February 2019 were collected. IORT applied 6 MeV or 9 MeV electron beams and a single irradiation was given to the margin. Kaplan-Meier method was used for the overall survival (OS) and disease-free survival (DFS) analysis; log-rank test was used for survival comparison among subgroups. The recurrence patterns and adverse reactions were recorded. Univariate and multivariate Cox proportional hazards models were used to analyze the factors influencing the OS and DFS.Results:A total of 64 patients were enrolled, with the median age [ M ( Q1, Q3)] of 57 years (49, 63) years. All patients included 55 males (85.9%) and 9 females (14.1%). The median dose of IORT was 15 Gy (range: 12-17 Gy). The median follow-up time was 83.3 (64.4, 91.9) months. The 1-year, 3-year, 5-year, 7-year, 10-year OS rates were 90.4%, 80.6%, 75.5%, 71.4% and 47.6%, respectively; the 1-year, 3-year, 5-year, 7-year,10-year DFS rates were 77.8%, 68.1%, 59.6%, 57.6% and 38.4%, respectively. Univariate Cox regression analysis indicated that preoperative serum alpha-fetoprotein (AFP) > 400 ng/ml was an independent risk factor for poor OS (> 400 ng/ml vs. ≤ 400 ng/ml: HR = 6.57, 95% CI: 2.16-19.96, P < 0.001), while not the independent influencing factor of poor DFS ( HR = 1.71, 95% CI: 0.65-4.52, P = 0.277). The age ≤ 60 years or not, gender, viral hepatitis or not, American Joint Committee on Cancer stage, tumor diameter (> 5 cm or not), tumor number, degree of tumor differentiation, microvascular invasion or not, microsatellite nodules or not, anatomical liver resection or not, and the dose of IORT ≤15 Gy or not were not the independent influencing factors of poor OS and DFS (all P > 0.05). Kaplan-Meier method analysis showed that patients with preoperative serum AFP ≤ 400 ng/ml (48 cases) had better OS compared with those with preoperative serum AFP>400 ng/ml (16 cases) (5-year OS rate: 84.8% vs. 44.9%; 7-year OS rate: 79.9% vs.37.4%), and the difference was statistically significant ( P = 0.002). There was no statistically significant difference in the DFS between the 2 groups ( P = 0.134). During the follow-up, 28 patients (43.8%) relapsed, including 17 cases (26.6%) of early recurrence and 11 cases (17.2%) of late recurrence. No marginal recurrence was observed. There were 22 cases (34.4%) of intrahepatic recurrence alone, 2 cases (3.1%) of extrahepatic recurrence and 4 cases (6.3%) of stimutaneous recurrence inside and outside the liver. The 1-, 3-, 5- and 7-year cumulative recurrence rates inside the liver were 19.0%, 27.2%, 37.4% and 39.3% respectively, and the cumulative recurrence rates outside the liver were 6.4%, 8.0%, 9.6% and 9.6% respectively. There were no adverse reactions above grade 3 in the entire group. There were no surgery-related deaths within 30 d after the operation, and no radiation-induced liver disease occurred. Conclusions:Narrow-margin IORT helps HCC patients receiving hepatectomy to achieve favorable long-term survival and adverse reactions are tolerable. It can be used as a safe and effective adjuvant therapy alternative.