Objective To investigate the effec of oscillating field stimulation on the proliferation of endogenous neural stem cell(ENSC)and recovery of motor function after spinal cord injury(SCI)in rats.Methods The SCI model was established by using the improved Allen's strike method and was randomly divided into experimental group and control group,with 60 rats in each group.The experimental group received an oscillating field implant as the intervention,while the control group was equipped with the same device but without activation.After successful modeling,the spinal cord of rats was removed on the 3rd,7th and 14th day,respectively.The proliferation of ENSC was detected by neuroepithelial stem cell protein(Nestin)/5-bromodeoxyuridine(BrdU)immunofluorescence,and the expression of Wnt-3a protein in spinal cord was detected by immunohistochemistry.At the 4th,6th and 8th week,the spinal cord was stained with osmic acid to observe the formation of myelin sheath in rats,and Basso-Beattle-Bresnahan(BBB)functional score of each group of rats was performed before sampling.Results The immunofluorescence results showed that Nestin/BrdU double positive cells in the injured area could be seen in both groups on the 3rd day after SCI,reaching a peak on the 7th day,and gradually decreased on the 14th day,and still maintained at a high level.At the same time,number of Nestin/BrdU double positive cells in experimental group rats at each time point was more than that in control group.The immunohistochemical results showed that the expression level of Wnt-3a protein in experimental group rats at each time point was higher than that in control group(P＜0.05).At the 6th and 8th weeks,the number of myelin sheaths in experimental group rats was significantly higher than that in control group(P＜0.05).At the 4 th,6th and 8th weeks,the BBB scores of rats in experimental group were significantly higher than those in control group(P＜0.05).Conclusion The oscillating field stimulation can induce the proliferation of ENSC after SCI by activating Wnt/β-catenin signaling pathway,promote myelin regeneration,and improve motor function of rats.

Objective To investigate the correlation between lipoprotein-associated phospholipase A2(Lp-PLA2),hemoglobin glycotion index(HGI)and the severity of type 2 diabetic retinopathy.Methods A total of 120 patients with type 2 diabetes mellitus who were hospitalized in the General Hospital of Hunan Medical University from January 2021 to June 2022 were selected and divided into type 2 diabetes without retinopathy group(NDR group,n=45),non-proliferative diabetic retinopathy group(NPDR group,n=45)and prolifera-tive diabetic retinopathy group(PDR group,n=30).In the same period,50 healthy subjects were randomly selected as control group.The clinical data,Lp-PLA2 and HGI levels of each group were collected and com-pared,and the correlation between different HGI and Lp-PLA2 levels and the severity of the disease was ana-lyzed.Results The levels of Lp-PLA2 and HGI in NDR,NPDR and PDR groups were higher than those in control group(P＜0.05).The proportion of patients with NPDR and PDR in high-Lp-PLA group(H-Lp-PLA2 group)and high-HGI group(H-HGI group)were higher than those in low-HGI group(L-HGI group)and low-Lp-PLA2 group(L-Lp-PLA2 group),and the differences were statistically significant(P＜0.05).Multiple Logistic regression analysis showed that Lp-PLA2 and HGI were risk factors for DR(P＜0.05),and were positively correlated with the severity of type 2 diabetic retinopathy(P＜0.05).Conclusion HGI and Lp-PLA2 are closely related to the severity of retinopathy in type 2 diabetes mellitus,and may be used as new indicators to predict the severity of the disease.

Objective:To explore the effects of neutrophil extracellular traps (NET) on the proliferation and migration of breast cancer cells and the related mechanisms.Methods:The peripheral blood neutrophils were separated using density gradient centrifugation, 100 nmol/L phorbol 12-myristate 13-acetate (PMA) was used to stimulate the formation of NET. Breast cancer cell line MDA-MB-231 was selected and treated with NET (NET group) or deoxyribonucleaseⅠ (DNase Ⅰ) digested NET (NET+DNase Ⅰ group),normal cultured MDA-MB-231 cells were used as the control group. In MDA-MB-231 cells of each group; cell proliferation ability was detected using CCK-8 assay; cell migration was detected by cell scratch assay and Transwell assay; real-time quantitative polymerase chain reaction (qRT-PCR) was used to detect the relative expressions of frizzled homolog 10 (FZD10), cyclin D1, E-cadherin and Vimentin immunofluorescence assay was performed to detect the formation of NET and the expressions of E-cadherin and Vimentin; Western blotting was used to detected the expression of FZD10-Wnt-β-catenin signaling pathway-related proteins.Results:Neutrophils were stimulated with PMA for 4 hours, and immunofluorescence assay results showed that NET exhibited fibrous reticular structure with high expression of citrullinated histone (citH3) and myeloperoxidase (MPO). The CCK-8 assay results showed that the absorbance values of cells in the control group, NET group and NET+DNase Ⅰ group were 1.25±0.06, 2.14±0.15 and 1.47±0.11, respectively, and the difference was statistically significant ( F = 80.60, P < 0.001). The results of scratch assay displayed that the percentage of wound closure in the control group, NET group and NET+DNase Ⅰ group were (36.2±1.3)%, (67.2±2.0)% and (46.3±3.2)%, respectively, and the difference was statistically significant ( F = 140.50, P < 0.001). Transwell assay indicated that the number of migrating cells in the control group, NET group and NET+DNase Ⅰ group were 317±18, 512±23 and 356±23, respectively, and the difference was statistically significant ( F = 75.39, P < 0.001). qRT-PCR assay revealed that the relative expressions of FZD10, cyclin D1 and Vimentin mRNA in the NET group were higher than those in the control group, while the relative expression of E-cadherin mRNA was lower than that in the control group, and the differences were statistically significant (all P < 0.001); the relative expressions of FZD10, cyclin D1 and Vimentin mRNA in the NET+DNase Ⅰ group were lower than those in the NET group, while the relative expression of E-cadherin mRNA was higher than that in the NET group, and the differences were statistically significant (all P < 0.001). Immunofluorescence assay indicated that the number of cells expressing Vimentin in the control group, NET group and NET+DNase Ⅰ group were 35±6, 86±13 and 51±6, respectively, and the difference was statistically significant ( F = 24.65, P = 0.001); the number of cells expressing E-cadherin were 31±4, 11±3 and 24±2, respectively, and the difference was statistically significant ( F = 38.36, P < 0.001). Western blotting results demonstrated that the relative expressions of FZD10, β-catenin and Vimentin proteins in the NET group were higher than those in the control group, while the relative expression of E-cadherin protein was lower than that in the control group; the relative expressions of FZD10, β-catenin and Vimentin proteins in the NET+DNase Ⅰ group were lower than those in the NET group, while the relative expression of E-cadherin protein was higher than that in the NET group. Conclusions:NET may promote the proliferation and migration of breast cancer cells by upregulating the expression of FZD10 and subsequently activating the Wnt-β-catenin signal pathway.

OBJECTIVE: To observe the effect of acupuncture on chondrocyte autophagy in rats of knee osteoarthritis (KOA) and explore its underlying mechanisms. METHODS: Forty male SPF-grade SD rats were randomized into a blank group, a model group, a suspension group, an acupuncture group, and a combined therapy group, 8 rats in each one. Except the blank group, KOA model was prepared by the injection with papain. The suspension exercise therapy (10 min each time, three times daily), acupuncture (at "Yanglingquan" [GB34], "Zusanli" [ST36], and "Dubi" [ST35] on the right side, 30 min each intervention, once daily) and the combined therapy (the suspension exercise therapy combined with acupuncture) were delivered in the suspension group, the acupuncture group and the combined therapy group, respectively. The intervention of each group was performed continuously for 6 days, and 4 consecutive weeks, at the interval of 1 day. Before and after intervention, Lequesne MG score was assessed in the rats. After intervention, HE staining was adopted to observe the cartilaginous tissue morphology of the right knee joints, and Mankin score was evaluated; the serum levels of interleukin (IL)-1β, IL-6, and tumor neurosis factor-α (TNF-α) were measured using ELISA; the real-time PCR was provided to determine the mRNA expression of collagen protein type Ⅱ(COL2), collagen protein type Ⅹ (COL10), phosphatidylinositol 3-kinase (PI3K), protein kinase B (Akt), mammalian target of rapamycin (mTOR), and autophagy-regulated protein (Beclin-1) in the cartilaginous tissue of the right knee joint; Western blot was employed to detect the protein expression of PI3K, phosphorylated PI3K (p-PI3K), Akt, phosphorylated Akt (p-Akt), mTOR, phosphorylated mTOR (p-mTOR) and Beclin-1 in the cartilaginous tissue of the right knee joint. RESULTS: Compared with the blank group, the rats in the model group showed the higher Lequesne MG score (P<0.01), thinner cartilage of the right knee, reduced chondrocytes and disordered arrangement, and higher Mankin score (P<0.01). Besides, in the model group, the serum levels of IL-1β, IL-6 and TNF-α were elevated (P<0.01), the mRNA expression of COL2 and Beclin-1 and the protein expression of Beclin-1 decreased (P<0.01), the mRNA expression of COL10, PI3K, Akt and mTOR, and the protein expression of p-PI3K, p-Akt and p-mTOR increased (P<0.01) in the cartilaginous tissue of the right knee joint. Compared with the model group, in the suspension group, the acupuncture group and the combined therapy group, the Lequesne MG scores were reduced (P<0.01), the cartilage of the right knee was thickened, the arrangement of chondrocytes was improved, and the Mankin scores were lower (P<0.01). Besides, in these intervention groups, the serum levels of IL-1β, IL-6 and TNF-α were reduced (P<0.01), the mRNA expression of COL2 and Beclin-1 and the protein expression of Beclin-1 increased (P<0.05, P<0.01), the mRNA expression of COL10, PI3K, Akt and mTOR, and the protein expression of p-PI3K, p-Akt and p-mTOR decreased (P<0.05, P<0.01) in the cartilaginous tissue of the right knee joint. When compared with the suspension group and the acupuncture group, in the combined therapy group, the Lequesne MG score was reduced (P<0.01), and the Mankin score was reduced (P<0.05, P<0.01). Besides, the serum levels of IL-1β, IL-6 and TNF-α were reduced (P<0.05), the mRNA expression of COL2 and Beclin-1 and the protein expression of Beclin-1 increased (P<0.05), the mRNA expression of COL10, PI3K, Akt and mTOR, and the protein expression of p-PI3K, p-Akt and p-mTOR decreased (P<0.05, P<0.01) in the cartilaginous tissue of the right knee joint. CONCLUSION Acupuncture can promote cartilage regeneration of knee joint and autophagy in KOA rats, alleviate inflammation, so as to retard cartilage degeneration, which may be possibly associated with the PI3K/Akt/mTOR signaling pathway.
Animals ; Male ; Acupuncture Therapy ; Proto-Oncogene Proteins c-akt/genetics* ; TOR Serine-Threonine Kinases/genetics* ; Osteoarthritis, Knee/physiopathology* ; Phosphatidylinositol 3-Kinases/genetics* ; Rats ; Signal Transduction ; Rats, Sprague-Dawley ; Chondrocytes/metabolism* ; Humans ; Autophagy ; Acupuncture Points

Objective To construct a predictive model for delayed neurological sequelae(DNS)following acute carbon monoxide poisoning(ACMP)and to verify its efficacy.Methods A retrospective analysis of the gen-eral data of 183 patients with ACMP was conducted.The factors influencing the occurrence of DNS were analyzed using a multivariate Logistic regression model.A corresponding predictive model was then established and its efficacy was verified.Results The multivariate logistic regression model showed that age,smoking history,severe poisoning,blood lactate,time from poisoning to hyperbaric oxygen therapy,and pulmonary infection were independent risk factors for DNS following ACMP(P<0.05).The area under the curve(AUC)of the model for predicting DNS in the development set was 0.933,with a sensitivity of 94.12%and specificity of 89.77%.In the validation set,the AUC was 0.906,with a sensitivity of 90.00%and specificity of 92.68%.The Hosmer-Lemeshow test showed that the predicted probabilities of DNS in both the development and validation sets were not significantly different from the actual probabilities(P>0.05).The predictive model achieved clinical net benefit within the risk threshold ranges of 0.11～0.98 for the development set and 0.12～0.92 for the validation set.Conclusions Age,smoking history,severe poisoning,blood lactate,time from poisoning to hyperbaric oxygen therapy,and pulmonary infec-tion are independent risk factors for DNS following ACMP.The corresponding predictive model has been verified to have good clinical efficacy.

Objective:To explore the effects of neutrophil extracellular traps (NET) on the proliferation and migration of breast cancer cells and the related mechanisms.Methods:The peripheral blood neutrophils were separated using density gradient centrifugation, 100 nmol/L phorbol 12-myristate 13-acetate (PMA) was used to stimulate the formation of NET. Breast cancer cell line MDA-MB-231 was selected and treated with NET (NET group) or deoxyribonucleaseⅠ (DNase Ⅰ) digested NET (NET+DNase Ⅰ group),normal cultured MDA-MB-231 cells were used as the control group. In MDA-MB-231 cells of each group; cell proliferation ability was detected using CCK-8 assay; cell migration was detected by cell scratch assay and Transwell assay; real-time quantitative polymerase chain reaction (qRT-PCR) was used to detect the relative expressions of frizzled homolog 10 (FZD10), cyclin D1, E-cadherin and Vimentin immunofluorescence assay was performed to detect the formation of NET and the expressions of E-cadherin and Vimentin; Western blotting was used to detected the expression of FZD10-Wnt-β-catenin signaling pathway-related proteins.Results:Neutrophils were stimulated with PMA for 4 hours, and immunofluorescence assay results showed that NET exhibited fibrous reticular structure with high expression of citrullinated histone (citH3) and myeloperoxidase (MPO). The CCK-8 assay results showed that the absorbance values of cells in the control group, NET group and NET+DNase Ⅰ group were 1.25±0.06, 2.14±0.15 and 1.47±0.11, respectively, and the difference was statistically significant ( F = 80.60, P < 0.001). The results of scratch assay displayed that the percentage of wound closure in the control group, NET group and NET+DNase Ⅰ group were (36.2±1.3)%, (67.2±2.0)% and (46.3±3.2)%, respectively, and the difference was statistically significant ( F = 140.50, P < 0.001). Transwell assay indicated that the number of migrating cells in the control group, NET group and NET+DNase Ⅰ group were 317±18, 512±23 and 356±23, respectively, and the difference was statistically significant ( F = 75.39, P < 0.001). qRT-PCR assay revealed that the relative expressions of FZD10, cyclin D1 and Vimentin mRNA in the NET group were higher than those in the control group, while the relative expression of E-cadherin mRNA was lower than that in the control group, and the differences were statistically significant (all P < 0.001); the relative expressions of FZD10, cyclin D1 and Vimentin mRNA in the NET+DNase Ⅰ group were lower than those in the NET group, while the relative expression of E-cadherin mRNA was higher than that in the NET group, and the differences were statistically significant (all P < 0.001). Immunofluorescence assay indicated that the number of cells expressing Vimentin in the control group, NET group and NET+DNase Ⅰ group were 35±6, 86±13 and 51±6, respectively, and the difference was statistically significant ( F = 24.65, P = 0.001); the number of cells expressing E-cadherin were 31±4, 11±3 and 24±2, respectively, and the difference was statistically significant ( F = 38.36, P < 0.001). Western blotting results demonstrated that the relative expressions of FZD10, β-catenin and Vimentin proteins in the NET group were higher than those in the control group, while the relative expression of E-cadherin protein was lower than that in the control group; the relative expressions of FZD10, β-catenin and Vimentin proteins in the NET+DNase Ⅰ group were lower than those in the NET group, while the relative expression of E-cadherin protein was higher than that in the NET group. Conclusions:NET may promote the proliferation and migration of breast cancer cells by upregulating the expression of FZD10 and subsequently activating the Wnt-β-catenin signal pathway.

Objective To investigate the effec of oscillating field stimulation on the proliferation of endogenous neural stem cell(ENSC)and recovery of motor function after spinal cord injury(SCI)in rats.Methods The SCI model was established by using the improved Allen's strike method and was randomly divided into experimental group and control group,with 60 rats in each group.The experimental group received an oscillating field implant as the intervention,while the control group was equipped with the same device but without activation.After successful modeling,the spinal cord of rats was removed on the 3rd,7th and 14th day,respectively.The proliferation of ENSC was detected by neuroepithelial stem cell protein(Nestin)/5-bromodeoxyuridine(BrdU)immunofluorescence,and the expression of Wnt-3a protein in spinal cord was detected by immunohistochemistry.At the 4th,6th and 8th week,the spinal cord was stained with osmic acid to observe the formation of myelin sheath in rats,and Basso-Beattle-Bresnahan(BBB)functional score of each group of rats was performed before sampling.Results The immunofluorescence results showed that Nestin/BrdU double positive cells in the injured area could be seen in both groups on the 3rd day after SCI,reaching a peak on the 7th day,and gradually decreased on the 14th day,and still maintained at a high level.At the same time,number of Nestin/BrdU double positive cells in experimental group rats at each time point was more than that in control group.The immunohistochemical results showed that the expression level of Wnt-3a protein in experimental group rats at each time point was higher than that in control group(P＜0.05).At the 6th and 8th weeks,the number of myelin sheaths in experimental group rats was significantly higher than that in control group(P＜0.05).At the 4 th,6th and 8th weeks,the BBB scores of rats in experimental group were significantly higher than those in control group(P＜0.05).Conclusion The oscillating field stimulation can induce the proliferation of ENSC after SCI by activating Wnt/β-catenin signaling pathway,promote myelin regeneration,and improve motor function of rats.

Objective To construct a predictive model for delayed neurological sequelae(DNS)following acute carbon monoxide poisoning(ACMP)and to verify its efficacy.Methods A retrospective analysis of the gen-eral data of 183 patients with ACMP was conducted.The factors influencing the occurrence of DNS were analyzed using a multivariate Logistic regression model.A corresponding predictive model was then established and its efficacy was verified.Results The multivariate logistic regression model showed that age,smoking history,severe poisoning,blood lactate,time from poisoning to hyperbaric oxygen therapy,and pulmonary infection were independent risk factors for DNS following ACMP(P<0.05).The area under the curve(AUC)of the model for predicting DNS in the development set was 0.933,with a sensitivity of 94.12%and specificity of 89.77%.In the validation set,the AUC was 0.906,with a sensitivity of 90.00%and specificity of 92.68%.The Hosmer-Lemeshow test showed that the predicted probabilities of DNS in both the development and validation sets were not significantly different from the actual probabilities(P>0.05).The predictive model achieved clinical net benefit within the risk threshold ranges of 0.11～0.98 for the development set and 0.12～0.92 for the validation set.Conclusions Age,smoking history,severe poisoning,blood lactate,time from poisoning to hyperbaric oxygen therapy,and pulmonary infec-tion are independent risk factors for DNS following ACMP.The corresponding predictive model has been verified to have good clinical efficacy.

ObjectiveTo assess the microstructural involvement of gray matter in recovered COVID-19 patients using Synthetic MRI. MethodsThis study was conducted in 29 recovered COVID-19 patients， including severe group （SG， n=11） and ordinary group （OG， n=18）. Healthy volunteers matched by age， sex， BMI and years of education were selected as a healthy control group （HC=23 cases）. Each subject underwent synthetic MRI to generate quantitative T₁ and T₂ maps， and the T₁ and T₂ maps were segmented into 90 regions of interest （ROIs） using automatic anatomical labeling （AAL） mapping. T₁ and T₂ values for each ROI were obtained by averaging all voxels within the ROIs. The T₁ and T₂ values of the 90 brain regions between the three groups were compared. ResultsRelative to HC， the SG had significantly higher T2 values in bilateral orbital superior frontal gyrus， bilateral parahippocampal gyrus， bilateral putamen， bilateral middle temporal gyrus， bilateral Inferior temporal gyrus， left orbital superior frontal gyrus， left orbital inferior frontal gyrus， left gyrus rectus， left anterior cingulate and paracingulate gyri， right median cingulate and paracingulate gyri， left posterior cingulate gyrus， and left supramarginal gyrus （P＜0.05）； Relative to OG， SG showed significantly increased T₂ values in the left rectus gyrus， left parahippocampal gyrus， bilateral middle temporal gyrus， and bilateral inferior temporal gyrus （P＜0.05）. Relative to HC， the T₁ values of SG were significantly increased in bilateral orbital superior frontal gyrus， left rectus gyrus， left anterior cingulate and paracingulate gyri， right posterior cingulate gyrus， left parahippocampal gyrus， left lingual gyrus， left putamen， left thalamus（P＜0.05）； Relative to OG， the T₁ values of SG were significantly higher in the right posterior cingulate gyrus， right calcarine fissure and surrounding cortex， and left putamen （P＜0.05）. ConclusionsEven after recovering from COVID-19， patients may still have persistent or delayed damage to their brain gray matter structure， which is correlated with the severity of the condition. SyMRI can serve as a sensitive tool to assess the extent of microstructural damage to the central nervous system， aiding in early diagnosis of the disease.

Patients with severe trauma require an extremely timely treatment and transfusion plays an irreplaceable role in the emergency treatment of such patients. An increasing number of evidence-based medicinal evidences and clinical practices suggest that patients with severe traumatic bleeding benefit from early transfusion of low-titer group O whole blood or hemostatic resuscitation with red blood cells, plasma and platelet of a balanced ratio. However, the current domestic mode of blood supply cannot fully meet the requirements of timely and effective blood transfusion for emergency treatment of patients with severe trauma in clinical practice. In order to solve the key problems in blood supply and blood transfusion strategies for emergency treatment of severe trauma, Branch of Clinical Transfusion Medicine of Chinese Medical Association, Group for Trauma Emergency Care and Multiple Injuries of Trauma Branch of Chinese Medical Association, Young Scholar Group of Disaster Medicine Branch of Chinese Medical Association organized domestic experts of blood transfusion medicine and trauma treatment to jointly formulate Chinese expert consensus on blood support mode and blood transfusion strategies for emergency treatment of severe trauma patients ( version 2024). Based on the evidence-based medical evidence and Delphi method of expert consultation and voting, 10 recommendations were put forward from two aspects of blood support mode and transfusion strategies, aiming to provide a reference for transfusion resuscitation in the emergency treatment of severe trauma and further improve the success rate of treatment of patients with severe trauma.