Hypoxic-ischemic (HI) brain damage poses a high risk of death or lifelong disability, yet effective treatments remain elusive. Here, we demonstrated that miR-100-5p levels in the lesioned cortex increased after HI insult in neonatal mice. Knockdown of miR-100-5p expression in the brain attenuated brain injury and promoted functional recovery, through inhibiting the cleaved-caspase-3 level, microglia activation, and the release of proinflammation cytokines following HI injury. Engineered extracellular vesicles (EVs) containing neuron-targeting rabies virus glycoprotein (RVG) and miR-100-5p antagonists (RVG-EVs-Antagomir) selectively targeted brain lesions and reduced miR-100-5p levels after intranasal delivery. Both pre- and post-HI administration showed therapeutic benefits. Mechanistically, we identified protein phosphatase 3 catalytic subunit alpha (Ppp3ca) as a novel candidate target gene of miR-100-5p, inhibiting c-Fos expression and neuronal apoptosis following HI insult. In conclusion, our non-invasive method using engineered EVs to deliver miR-100-5p antagomirs to the brain significantly improves functional recovery after HI injury by targeting Ppp3ca to suppress neuronal apoptosis.
Animals ; MicroRNAs/metabolism* ; Extracellular Vesicles/metabolism* ; Mice ; Recovery of Function/physiology* ; Hypoxia-Ischemia, Brain/therapy* ; Mice, Inbred C57BL ; Antagomirs/administration & dosage* ; Male ; Animals, Newborn ; Apoptosis/drug effects* ; Brain Injuries/metabolism* ; Glycoproteins ; Peptide Fragments ; Viral Proteins

Objective:To investigate the effect of aerobic exercise with acupuncture needles on the learning and memory ability in rats subjected to middle cerebral artery occlusion/reperfusion(MCAO/R)and to explore the underly-ing mechanisms.Method:Male SD rats were randomly divided into four groups:sham,model,aerobic exercise with acupunc-ture needles,and inhibitor groups(n=12 per group).In the sham group,only the left common carotid artery,in-ternal carotid ar tery,and external carotid artery were exposed without inducing ischemia,while the other three groups underwent MCAO/R modeling.To establish an MCAO/R model with reduced NLRP3 expression,rats in the inhibitor group received an intraperitoneal injection of MCC950 60 minutes before modeling.On the first day of post-modeling,the Zea-Longa score and T2 WI were used to screen the MCAO/R rats.From the second day,rats in the aerobic exercise with acupuncture needles and the inhibitor groups were underwent a 14-day intervention.The acupoints were selected for"Shenting"(GV 24)and"Baihui"(GV 20),with sparse waveform at a frequency 2/10Hz and a current intensity of 1-3mA.Simultaneously,the rats performed tread-mill exercise,with speeds of 4m/min(day 1-3),6m/min(day 4-7),and 8m/min(day 8-14)for 30min/day.Afterthe intervention,neurological function was reassessed,and the new object recognition test was used to measure the learning and memory ability,and the pathological changes of the hippocampus were observed by HE staining.The protein expressions of NLRP 3,GSDMD,and Caspase-1 were detected by Western Blot and immunofluorescence,ELISA detected the contents of ILL-1β and IL-18,the expression and localization of IBA 1 and NLRP 3 were detected by double immunofluorescent staining.Result:The hippocampal neurons in the ischemic side of the rats were loosely and disorderly arranged in the model group.Compared to the sham group,the model group showed significantly impaired neurological func-tion and novel object recognition ability(P＜0.01),with significant increased expression of NLRP 3,GSDMD,and Caspase-1(P＜0.01),and elevated ILL-lβ and IL-18 contents(P＜0.01).In addition,IBA 1 and NLRP 3 fluo-rescence expression were enhanced(P＜0.01),and many IBA 1+/NLRP 3+double-positive cells were observed un-der the microscope(P＜0.01).Compared with the model group,the aerobic exercise with acupuncture needles group showed significant improvements in neurological function and the new object recognition index(P＜0.05,P＜0.01),reduced hippocampal pathological injury,decreased expression of NLRP 3,GSDMD,and Caspase-1 d(P＜0.01)and lower IL-lβ and IL-18 levels(P＜0.05);the activation of microglia and the expression of IBA 1,NL-RP 3 and the number of IBA 1+/NLRP 3+double-positive cells decreased(P＜0.01).The inhibitor group demon-strated even greater improvements in neural function and new object recognition performance than the aerobic exercise with acupuncture needles group(P＜0.05,P＜0.01),with further reductions in NLRP 3,GSDMD,Caspase-1 protein expression(P＜0.01)and lower level of IL-1β,IL-18(P＜0.01,P＜0.05);the microglia activation level,the number,arrangement and morphology of nerve cells were closely resembled those in the sham group,and there was minimal NLRP 3 microglia observed under the microscope(P＜0.01).Conclusion:Aerobic exercise with acupuncture needles at"Shenting"(GV 24)and"Baihui"(GV 20)points could improve the learning and memory function in MACO/R rats by inhibiting microglia pyroptosis and activa-tion,thereby reducing neuroinflammation.

ObjectiveTo investigate the influencing factors for recompensation in patients with decompensated hepatitis C cirrhosis， and to establish a predictive model. MethodsA total of 217 patients who were diagnosed with decompensated hepatitis C cirrhosis and were admitted to The Third People’s Hospital of Kunming l from January， 2019 to December， 2022 were enrolled， among whom 63 patients who were readmitted within at least 1 year and had no portal hypertension-related complications were enrolled as recompensation group， and 154 patients without recompensation were enrolled as control group. Related clinical data were collected， and univariate and multivariate analyses were performed for the factors that may affect the occurrence of recompensation. The independent-samples t test was used for comparison of normally distributed measurement data between two groups， and the Mann-Whitney U test was used for comparison of non-normally distributed measurement data between two groups； the chi-square test or the Fisher’s exact test was used for comparison of categorical data between two groups. A binary Logistic regression analysis was used to investigate the influencing factors for recompensation in patients with decompensated hepatitis C cirrhosis， and the receiver operating characteristic （ROC） curve was used to assess the predictive performance of the model. ResultsAmong the 217 patients with decompensated hepatitis C cirrhosis， 63 （29.03%） had recompensation. There were significant differences between the recompensation group and the control group in HIV history （χ²=4.566， P=0.034）， history of partial splenic embolism （χ²=6.687， P=0.014）， Child-Pugh classification （χ²=11.978， P=0.003）， grade of ascites （χ²=14.229， P<0.001）， albumin （t=4.063， P<0.001）， prealbumin （Z=-3.077， P=0.002）， high-density lipoprotein （t=2.854， P=0.011）， high-sensitivity C-reactive protein （Z=-2.447， P=0.014）， prothrombin time （Z=-2.441， P=0.015）， carcinoembryonic antigen （Z=-2.113， P=0.035）， alpha-fetoprotein （AFP） （Z=-2.063， P=0.039）， CA125 （Z=-2.270， P=0.023）， TT3 （Z=-3.304， P<0.001）， TT4 （Z=-2.221， P=0.026）， CD45⁺ （Z=-2.278， P=0.023）， interleukin-5 （Z=-2.845， P=0.004）， tumor necrosis factor-α （Z=-2.176， P=0.030）， and portal vein width （Z=-5.283， P=0.005）. The multivariate analysis showed that history of partial splenic embolism （odds ratio ［OR］=3.064， P=0.049）， HIV history （OR=0.195， P=0.027）， a small amount of ascites （OR=3.390， P=0.017）， AFP （OR=1.003， P=0.004）， and portal vein width （OR=0.600， P<0.001） were independent influencing factors for the occurrence of recompensation in patients with decompensated hepatitis C cirrhosis. The ROC curve analysis showed that HIV history， grade of ascites， history of partial splenic embolism， AFP， portal vein width， and the combined predictive model of these indices had an area under the ROC curve of 0.556， 0.641， 0.560， 0.589， 0.745， and 0.817， respectively. ConclusionFor patients with decompensated hepatitis C cirrhosis， those with a history of partial splenic embolism， a small amount of ascites， and an increase in AFP level are more likely to experience recompensation， while those with a history of HIV and an increase in portal vein width are less likely to experience recompensation.

Objective:To investigate the effect of aerobic exercise with acupuncture needles on the learning and memory ability in rats subjected to middle cerebral artery occlusion/reperfusion(MCAO/R)and to explore the underly-ing mechanisms.Method:Male SD rats were randomly divided into four groups:sham,model,aerobic exercise with acupunc-ture needles,and inhibitor groups(n=12 per group).In the sham group,only the left common carotid artery,in-ternal carotid ar tery,and external carotid artery were exposed without inducing ischemia,while the other three groups underwent MCAO/R modeling.To establish an MCAO/R model with reduced NLRP3 expression,rats in the inhibitor group received an intraperitoneal injection of MCC950 60 minutes before modeling.On the first day of post-modeling,the Zea-Longa score and T2 WI were used to screen the MCAO/R rats.From the second day,rats in the aerobic exercise with acupuncture needles and the inhibitor groups were underwent a 14-day intervention.The acupoints were selected for"Shenting"(GV 24)and"Baihui"(GV 20),with sparse waveform at a frequency 2/10Hz and a current intensity of 1-3mA.Simultaneously,the rats performed tread-mill exercise,with speeds of 4m/min(day 1-3),6m/min(day 4-7),and 8m/min(day 8-14)for 30min/day.Afterthe intervention,neurological function was reassessed,and the new object recognition test was used to measure the learning and memory ability,and the pathological changes of the hippocampus were observed by HE staining.The protein expressions of NLRP 3,GSDMD,and Caspase-1 were detected by Western Blot and immunofluorescence,ELISA detected the contents of ILL-1β and IL-18,the expression and localization of IBA 1 and NLRP 3 were detected by double immunofluorescent staining.Result:The hippocampal neurons in the ischemic side of the rats were loosely and disorderly arranged in the model group.Compared to the sham group,the model group showed significantly impaired neurological func-tion and novel object recognition ability(P＜0.01),with significant increased expression of NLRP 3,GSDMD,and Caspase-1(P＜0.01),and elevated ILL-lβ and IL-18 contents(P＜0.01).In addition,IBA 1 and NLRP 3 fluo-rescence expression were enhanced(P＜0.01),and many IBA 1+/NLRP 3+double-positive cells were observed un-der the microscope(P＜0.01).Compared with the model group,the aerobic exercise with acupuncture needles group showed significant improvements in neurological function and the new object recognition index(P＜0.05,P＜0.01),reduced hippocampal pathological injury,decreased expression of NLRP 3,GSDMD,and Caspase-1 d(P＜0.01)and lower IL-lβ and IL-18 levels(P＜0.05);the activation of microglia and the expression of IBA 1,NL-RP 3 and the number of IBA 1+/NLRP 3+double-positive cells decreased(P＜0.01).The inhibitor group demon-strated even greater improvements in neural function and new object recognition performance than the aerobic exercise with acupuncture needles group(P＜0.05,P＜0.01),with further reductions in NLRP 3,GSDMD,Caspase-1 protein expression(P＜0.01)and lower level of IL-1β,IL-18(P＜0.01,P＜0.05);the microglia activation level,the number,arrangement and morphology of nerve cells were closely resembled those in the sham group,and there was minimal NLRP 3 microglia observed under the microscope(P＜0.01).Conclusion:Aerobic exercise with acupuncture needles at"Shenting"(GV 24)and"Baihui"(GV 20)points could improve the learning and memory function in MACO/R rats by inhibiting microglia pyroptosis and activa-tion,thereby reducing neuroinflammation.

Lactic acid bacteria(LAB)were isolated and cultivated from the intestinal contents of rabbits by MRS-CaCO3 solid medium.Identification was achieved through morphological observa-tion,Gram staining,physiological and biochemical characterisation,16S rDNA sequence analysis,and ERIC-PCR analysis.Strains displaying typical Lactobacilli characteristics were exanimated for their biological characteristics,resistance properties,adherence capacity in vitro,colonization abili-ty in vivo,and safety profile.In this study,a total of four strains of Lactobacillus reuteri were iso-lated from rabbits,all of which exhibited typical biological characteristics of LAB.These strains demonstrated inhibitory effects on common pathogenic bacteria in the gastrointestinal tract,with the primary inhibitory substance being bacteriocin.Furthermore,they showed sensitivity to chlor-amphenicol,rifampicin,and erythromycin,and displayed a degree of tolerance to gastrointestinal conditions and high temperature.These stains were capable of successful colonization in rabbits with a higher degree of safety.This study lays a foundation for the development of LAB prepara-tions for the prevention and treatment of rabbit intestinal diseases.

Objective:To construct of a computed tomography (CT) based radiomics model for predicting the prognosis of patients with gastric neuroendocrine neoplasm (GNEN) and inves-tigate its application value.Methods:The retrospective cohort study was conducted. The clinico-pathological data of 182 patients with GNEN who were admitted to 2 medical centers, including the First Affiliated Hospital of Zhengzhou University of 124 cases and the Affiliated Cancer Hospital of Zhengzhou University of 58 cases, from August 2011 to December 2020 were collected. There were 130 males and 52 females, aged 64(range, 56-70)years. Based on random number table, all 182 patients were divided into the training dataset of 128 cases and the validation dataset of 54 cases with a ratio of 7:3. All patients underwent enhanced CT examination. Observation indicators: (1) construction and validation of the radiomics prediction model; (2) analysis of prognostic factors for patients with GNEN in the training dataset; (3) construction and evaluation of the prediction model for prognosis of patients with GNEN. Measurement data with skewed distribution were represented as M(range), and comparison between groups was conducted using the Mann-Whitney U test. Count data were described as absolute numbers, and the chi-square test, corrected chi-square test or Fisher exact probability were used for comparison between groups. The Kaplan-Meier method was used to calculate survival rate and draw survival curve, and the Log-rank test was used for survival analysis. The COX regression model was used for univariate and multivariate analyses. The R software (version 4.0.3) glmnet software package was used for least absolute shrinkage and selection operator (LASSO)-COX regression analysis. The rms software (version 4.0.3) was used to generate nomogram and calibration curve. The Hmisc software (version 4.0.3) was used to calculate C-index values. The dca.R software (version 4.0.3) was used for decision curve analysis. Results:(1) Construction and valida-tion of the radiomics prediction model. One thousand seven hundred and eighty-one radiomics features were finally extracted from the 182 patients. Based on the feature selection using intra-group correlation coefficient >0.75, and the reduce dimensionality using LASSO-COX regression analysis, 14 non zero coefficient radiomics features were finally selected from the 1 781 radiomics features. The radiomics prediction model was constructed based on the radiomics score (R-score) of these non zero coefficient radiomics features. According to the best cutoff value of the R-score as -0.494, 128 patients in the training dataset were divided into 64 cases with high risk and 64 cases with low risk, 54 patients in the validation dataset were divided into 35 cases with high risk and 19 cases with low risk. The area under curve (AUC) of radiomics prediction model in predicting 18-, 24-, 30-month overall survival rate of patients in the training dataset was 0.83[95% confidence interval ( CI ) as 0.76-0.87, P<0.05], 0.84(95% CI as 0.73-0.91, P<0.05), 0.91(95% CI as 0.78-0.95, P<0.05), respectively. The AUC of radiomics prediction model in predicting 18-, 24-, 30-month overall survival rate of patients in the validation dataset was 0.84(95% CI as 0.75-0.92, P<0.05), 0.84 (95% CI as 0.73-0.91, P<0.05), 0.86(95% CI as 0.82-0.94, P<0.05), respectively. (2) Analysis of prognostic factors for patients with GNEN in the training dataset. Results of multivariate analysis showed gender, age, treatment method, tumor boundary, tumor T staging, tumor N staging, tumor M staging, Ki-67 index, CD56 expression were independent factors influencing prognosis of patients with GNEN in the training dataset ( P<0.05). (3) Construction and evaluation of the prediction model for prognosis of patients with GNEN. The clinical prediction model was constructed based on the independent factors influen-cing prognosis of patients with GNEN including gender, age, treatment method, tumor boundary, tumor T staging, tumor N staging, tumor M staging, Ki-67 index, CD56 expression. The C-index value of clinical prediction model in the training dataset and the validation dataset was 0.86 (95% CI as 0.82-0.90) and 0.80(95% CI as 0.72-0.87), respectively. The C-index value of radiomics prediction model in the training dataset and the validation dataset was 0.80 (95% CI as 0.74-0.86, P<0.05) and 0.75(95% CI as 0.66-0.84, P<0.05), respectively. The C-index value of clinical-radiomics combined prediction model in the training dataset and the validation dataset was 0.88(95% CI as 0.85-0.92) and 0.83 (95% CI as 0.77-0.89), respectively. Results of calibration curve show that clinical prediction model, radiomics prediction model and clinical-radiomics combined prediction model had good predictive ability. Results of decision curve show that the clinical-radiomics combined prediction model is superior to the clinical prediction model, radiomics prediction model in evaluating the prognosis of patients with GNEN. Conclusions:The predection model for predicting the prognosis of patients with GNEN is constructed based on 14 radiomics features after selecting. The prediction model can predict the prognosis of patients with GNEN well, and the clinical-radiomics combined prediction model has a better prediction efficiency.

【Objective】 To investigate the correlation between the titer of anti-A or anti-B antibodies before and after the absorption of IgG anti-AB antibodies in the serum of type O mothers with ABO hemolytic disease of fetus and newborn (ABO-HDFN) and the total bilirubin in the serum of the children. 【Methods】 Serum samples from 119 children diagnosed with ABO-HDFN and their mothers sent to the Beijing Red Cross Blood Center from January to December 2020 were selected, and clinical data of the children were collected. Three hemolysis tests and serum total bilirubin (TBIL) determination were conducted on the children. IgG anti-A or anti-B antibody titers were tested before and after the mother′s serum absorbed IgG anti-AB antibodies. Statistical analysis was conducted on the IgG antibody titers and the TBIL results of the children. The differences in TBIL results corresponding to different IgG antibody titers were compared. The Spearman test was used to analyze the correlation between the IgG anti-A or -B antibody titers and TBIL results before and after the absorption of IgG anti-AB antibodies. 【Results】 There were differences in the TBIL results corresponding to IgG anti-A or anti-B titers at different levels in the serum of type O mothers after absorption by IgG anti-AB antibodies (F=8.401, 19.622, P<0.05). There was no significant correlation between the IgG anti-A or anti-B titers of maternal serum before absorption by IgG anti-AB antibodies and neonatal TBIL results (r=0.181, 0.248, P>0.05). The IgG anti-A or anti-B titers of maternal serum absorbed by IgG anti-AB antibodies were positively correlated with neonatal TBIL results (r=0.487, 0.629, P<0.05). 【Conclusion】 There is a positive correlation between the titer of IgG anti-A or anti-B antibodies in the serum of type O mothers after absorbing IgG anti-AB antibodies and the TBIL results of ABO-HDFN children. The trend of increased total bilirubin in newborn serum ban be accurately predicted by detecting the titer level of IgG anti-A or anti-B antibodies in the serum of mothers after absorbing IgG anti-AB antibodies.

【Objective】 To investigate the inactivation function of 25Gy X-ray irradiation on apheresis platelets’ lymphocytes and its effect on the quantity of apheresis platelets(AP). 【Methods】 Twenty healthy voluntary AP donors from January to May 2021 in our center were selected, and 2 bags of AP were donated by each of them. The APs were divided into two groups to undergo X-ray and γ-ray irradiation for 10 min. Lymphocytes were separated from AP samples, before and after irradiation, by lymphocyte separation solution to analyze and compare the effect of X-ray and γ-ray on lymphocyte proliferation. The CD41b, CD62p, blood routine and pH of APs before and 1-day/3-day after irradiation were detected. SPSS statistical software was used to analyze and compare the differences between groups by independent sample t-test. 【Results】 After 25Gy X-ray and γ-ray irradiation, the inhibition rates of lymphocytes were (98.034±1.778)% and (97.882±1.915)%, respectively.Compared with the γ irradiation group, the difference of Plt, PDW, MPV, P-LCR, PCT, pH, CD41b and CD62p between 1-day and 3-day group after 25Gy X-ray irradiation showed not statistically significance (P>0.05). 【Conclusion】 25Gy X-ray irradiation can effectively inactivate lymphocytes in APs, and the radiation effect was equivalent with γ-ray; at the same time, there was no significant influence on the quantity of APs.

BACKGROUND:Adipose-derived mesenchymal stem cells are gaining widespread interest in the Achil es tendon tissue engineering and regeneration, and an enabling environment (oxygen concentration) for cellinduction and differentiation is particularly necessary.
OBJECTIVE:To co-culture adipose-derived mesenchymal stem cells with primary tenocytes in standard culture condition (20%O 2 tension) and in an atmosphere of reduced oxygen (2%O 2 tension) in order to determine whether the two conditions differ in their effect on tenogenic differentiation.
METHODS:Tenocytes were isolated via serial expansion in culture from several Sprague-Dawley rats’ Achil es tendons. Adipose-derived mesenchymal stem cells were purchased. After one passage, adipose-derived mesenchymal stem cells were indirectly co-cultured with tenocytes in standard culture condition (20%O 2 tension) and in an atmosphere of reduced oxygen (2%O 2 tension). Col agen 1, col agen 3, Tenomodulin, Thrombospondin-4, Scleraxis levels were compared for each culture condition at 7, 14 and 21 days fol owing co-culturing. Immunofluorescence staining was performed to evaluate production of col agen 1 and Thrombospondin-4.
RESULTS AND CONCLUSION:Fol owing indirect co-culturing, hypoxic differentiated adipose-derived mesenchymal stem cells expressed higher levels of tendon-related genes and proteins than normoxic controls, which suggest that oxygen levels can significantly affect tenogenic differentiation, and hypoxia is advantageous for efficient differentiation of adipose-derived mesenchymal stem cells in vitro for tendon tissue engineering.