Objective To prepare neutralizing serum against rubella virus(RuV) using mRNA technology and identify it, so as to apply it to the identification and virus titer detection of measles, mumps and rubella vaccine(MMR).Methods The mRNA lipid nanoparticle(LNP) vaccine expressing RuV BRDⅡ vaccine strain and 2B genotype wild strain E1 protein was constructed, and its expression in 293T cells was verified by Western blot and indirect immunofluorescence. Guinea pigs were immunized with two kinds of mRNA-LNP vaccines and live attenuated rubella vaccine separately to prepare antiserum. The indirect ELISA method was used to detect the specific IgG antibody titer of RuV E1 protein in the serum, and the micro-neutralization method was applied to determine the neutralizing antibody titer. Furthermore, the cross-interference effect of serum on mumpsvirusandmeaslesviruswasevaluated.Results TheparticlesizesofthetwomRNA-LNPvaccineswere71.69 and70.77 nm respectively, with the encapsulation efficiency of more than 85%, and the E1 protein could be effectively expressed in 293T cells. The E1 protein-specific IgG antibody titers and neutralizing antibody titers produced by guinea pigs immunized with the two mRNA-LNP vaccines were all significantly higher than those of guinea pigs immunized with the live attenuated rubella vaccine(F = 20. 38-37. 94 and 10. 63-21. 59, P < 0. 001 and < 0. 05,respectively). Additionally, the serum only specifically neutralized RuV, and had no cross-interference with measles virus and mumps virus.Conclusion Based on mRNA technology, a high-titer and high-specificity anti-RuV neutralizing serum was successfully prepared, which is suitable for the quality verification of MMR vaccines and provides a new strategy for the development of rubella vaccines.

Objective To explore the predictive value of maximum standardized uptake value(SUVmax)and mean standardized uptake value(SUVmean)parameters in 18F-prostate specific membrane antigen-1007 positron emission tomography/computed tomography(18F-PSMA-1007 PET/CT)for the diagnosis and disease progression of prostate cancer(PCa).Methods Eighty patients with suspected PCa who underwent 18F-PSMA-1007 PET/CT examination in the hospital were selected and divided into the PCa group(49 cases)and the non-PCa group(31 cases)according to the biopsy pathological results.The 18F-PSMA-1007 PET/CT parameters were collected in the two groups of patients at admission,including SUVmax and SUVmean.The prostate specific antigen(PSA)and prostate cancer histological grade(GLEASON)score were also collected in patients of the PCa group at admission and 3 months after admission.The diagnostic efficiency of 18F-PSMA-1007 PET/CT parameters on PCa was analyzed by receiver operating characteristic(ROC)curve.The correlation between 18F-PSMA-1007 PET/CT parameters of patients in the PCa group at admission and PSA,GLEASON score was analyzed by Pearson method.Results The SUVmax and SUVmean in the PCa group were higher than those in the non-PCa group(P＜0.05).ROC curve analysis showed that the areas under the curves(AUCs)of SUVmax and SUVmean were 0.887 and 0.843 respectively,the sensitivities were 80.65%and 83.87%,and the specificities were 87.76%and 73.47%respectively.The PSA and GLEASON score were lower 3 months after admission than those at admission in the PCa group(P＜0.05).Pearson correlation analysis showed that the SUVmax and SUVmean of patients in the PCa group at admission were positively correlated with PSA and GLEASON score at admission and 3 months after admission(P＜0.05).Conclusion SUVmax and SUVmean parameters of 18F-PSMA-1007 PET/CT have high predictive value for the diagnosis and disease progression of PCa.

Nocardia is a Gram-positive pathogen responsible for causing dairy mastitis,which leads to purulent granulomatous lesions in mammary tissue and can significantly impact the dairy indus-try,resulting in substantial economic losses.To develop a rapid and accurate diagnostic method for detecting Nocardia of bovine origin,a conserved sequence of the 16S rRNA gene from Nocardia was selected from the NCBI database.Based on this sequence,a pair of primers and a TaqMan fluo-rescent quantitative probe were designed.The validation of the TaqMan fluorescence quantitative PCR(qPCR)method found in this study showed that the Ctvalue had a good linear relationship with recombinant plasmid concentrations ranging from 1×1010 to 1×102 copies/μL,with a regres-sion equation of y=-3.536x+43.78,a correlation coefficient(R2)of 0.997 5,a slope of-3.536,and an amplification efficiency(E)of 91％(where 90％＜E＜110％).The specificity was strong,with no cross-reactions with other pathogens.The standard curve had a high sensitivity with a low-er detection limit of 1 × 102 copies/μL,it was 100-fold higher than conventional PCR.The repeatability of the standard curve was also good.Both intra-and inter-group coefficients of varia-tion were below 2％.Using this method,234 milk samples and 80 environmental samples were tested using this method,respectively,with a positive detection rate of 27.07％,whereas conven-tional PCR had a positive detection rate of 19.43％,indicating that this method was more sensitive compared to conventional PCR.The fluorescent quantitative PCR detection method established in this study provides an effective means for the clinical detection of Nocardia in dairy cows.

Nocardia is a Gram-positive pathogen responsible for causing dairy mastitis,which leads to purulent granulomatous lesions in mammary tissue and can significantly impact the dairy indus-try,resulting in substantial economic losses.To develop a rapid and accurate diagnostic method for detecting Nocardia of bovine origin,a conserved sequence of the 16S rRNA gene from Nocardia was selected from the NCBI database.Based on this sequence,a pair of primers and a TaqMan fluo-rescent quantitative probe were designed.The validation of the TaqMan fluorescence quantitative PCR(qPCR)method found in this study showed that the Ctvalue had a good linear relationship with recombinant plasmid concentrations ranging from 1×1010 to 1×102 copies/μL,with a regres-sion equation of y=-3.536x+43.78,a correlation coefficient(R2)of 0.997 5,a slope of-3.536,and an amplification efficiency(E)of 91％(where 90％＜E＜110％).The specificity was strong,with no cross-reactions with other pathogens.The standard curve had a high sensitivity with a low-er detection limit of 1 × 102 copies/μL,it was 100-fold higher than conventional PCR.The repeatability of the standard curve was also good.Both intra-and inter-group coefficients of varia-tion were below 2％.Using this method,234 milk samples and 80 environmental samples were tested using this method,respectively,with a positive detection rate of 27.07％,whereas conven-tional PCR had a positive detection rate of 19.43％,indicating that this method was more sensitive compared to conventional PCR.The fluorescent quantitative PCR detection method established in this study provides an effective means for the clinical detection of Nocardia in dairy cows.

Objective To explore the predictive value of maximum standardized uptake value(SUVmax)and mean standardized uptake value(SUVmean)parameters in 18F-prostate specific membrane antigen-1007 positron emission tomography/computed tomography(18F-PSMA-1007 PET/CT)for the diagnosis and disease progression of prostate cancer(PCa).Methods Eighty patients with suspected PCa who underwent 18F-PSMA-1007 PET/CT examination in the hospital were selected and divided into the PCa group(49 cases)and the non-PCa group(31 cases)according to the biopsy pathological results.The 18F-PSMA-1007 PET/CT parameters were collected in the two groups of patients at admission,including SUVmax and SUVmean.The prostate specific antigen(PSA)and prostate cancer histological grade(GLEASON)score were also collected in patients of the PCa group at admission and 3 months after admission.The diagnostic efficiency of 18F-PSMA-1007 PET/CT parameters on PCa was analyzed by receiver operating characteristic(ROC)curve.The correlation between 18F-PSMA-1007 PET/CT parameters of patients in the PCa group at admission and PSA,GLEASON score was analyzed by Pearson method.Results The SUVmax and SUVmean in the PCa group were higher than those in the non-PCa group(P＜0.05).ROC curve analysis showed that the areas under the curves(AUCs)of SUVmax and SUVmean were 0.887 and 0.843 respectively,the sensitivities were 80.65%and 83.87%,and the specificities were 87.76%and 73.47%respectively.The PSA and GLEASON score were lower 3 months after admission than those at admission in the PCa group(P＜0.05).Pearson correlation analysis showed that the SUVmax and SUVmean of patients in the PCa group at admission were positively correlated with PSA and GLEASON score at admission and 3 months after admission(P＜0.05).Conclusion SUVmax and SUVmean parameters of 18F-PSMA-1007 PET/CT have high predictive value for the diagnosis and disease progression of PCa.

Due to the drawbacks of low specificity and high risk of side effects of traditional anti-tumor treatments in clinical practice,novel anti-tumor immunotherapy has received attention and has been gradually applied.Tumor immunotherapy is to enhance the anti-tumor immune response by regulating the body's immune system in order to achieve control and killing of tumors.Tumor immunotherapies include immune checkpoint blockade therapy,over-the-counter cellular immunotherapy and tumor vaccines.Among them,the tumor vaccine stimulates the immune system to produce specific immune cells or antibodies by delivering tumor cell-specific antigens thereby eliminating the tumor cells for the purpose of treating the tumor.In recent years,The field of mRNA vaccines is developing rapidly,and the required mRNA in the synthesis and preparation of the process has been developed and matured,laying a good foundation for the research of tumor mRNA vaccine.Because of the fact that mRNA is easily degraded and cannot enter the cell autonomously,this vaccine requires a suitable delivery vehicle to be successfully taken up by the cell to be effective.Therefore,the development of mRNA vaccine delivery systems has become critical for their better utilization,which is also an important part of whether mRNA vaccines can be developed and utilized for the clinical stage in the field of tumor therapy.This paper briefly introduced the immunotherapeutic methods for tumors,types of tumor vaccines and the mechanism of action and preparation methods of tumor mRNA vaccines,reviewed the research progress and related applications of mRNA vaccines and their common delivery systems in immunotherapy for tumor treatment,and summarized the tumor mRNA vaccines that entered into the phase of clinical trials with the aim of providing assistance for the research of mRNA vaccines for tumors in the future.

Objective:To investigate the risk factors affecting the delay of recovery in patients under general anesthesia.Methods:Patients who underwent radical mastectomy for breast cancer in our hospital from Jul. 2020 to Aug. 2022 were selected as the research objects, and the effective data of 80 patients were obtained after screening. The patients were divided into the non-delayed recovery group (54 cases) and the delayed recovery group (26 cases). The general conditions and perioperative data of the two groups were compared, and the high-risk factors affecting delayed recovery from anesthesia were analyzed using the Logistic hazard proportional regression model. Results:In general, there were no significant differences in body mass index, hypertension, diabetes, tumor size, tumor subtype, tumor location, or tumor stage between the non-delayed awakening group and the delayed awakening group (all P>0.05). The average age of patients in the wake-up delay group was (53.28±11.01), the proportion of anemia was 42.30% (11/26), and the proportion of ASA Ⅱ patients was 76.92% (20/26) compared with the average age of the non-wake-up delay group (46.89±6.91) ( t=3.17, P=0.002), the proportion of anemia was 20.37% (11/54) ( χ2=3.17, P=0.040), the proportion of ASA Ⅱ patients was 27.78% (15/54) ( χ2=17.22, P<0.001), which was significantly increased. In the perioperative data, there was no statistical significance in the intraoperative combined epidural anesthesia between the non-delayed recovery group and the delayed recovery group (all P>0.05). The data of the patients in the delayed recovery group, including average intraoperative blood loss (234.14±32.28), operation time (229.47±29.84), anesthesia time (246.14±35.64) and intraoperative compound sevoflurane inhalation accounted for 69.23% (18/26) ,which were significantly increased compared with the data of non-delayed recovery group following, including the average intraoperative blood loss (215.48±29.54) ( t=2.57, P=0.012), operation time (206.35±27.41) ( t=3.43, P=0.001), anesthesia time (215.61±28.54) ( t=4.13, P<0.001), intraoperative compound sevoflurane inhalation (44.44%, 24/54). Through Logistic hazard ratio regression analysis, it was found that age ( OR=1.15, 95% CI: 1.05-1.30, P=0.008), ASA Ⅱ grade ( OR=9.49, 95% CI: 2.05-60.94, P=0.008), intraoperative bleeding volume ( OR=1.04, 95% CI: 1.01-1.08, P=0.012), operation time ( OR=1.05, 95% CI: 1.01-1.08, P=0.009), and anesthesia time ( OR=1.04, 95% CI: 1.02-1.07, P=0.004) were high-risk factors affecting the delayed recovery from anesthesia. Conclusion:Increasing age, high grade of ASA, heavy intraoperative blood loss, long operation time and anesthesia time are independent risk factors affecting delayed recovery from anesthesia.

Objective:To explore the effect of smoking on the wound healing of stage 4 pressure ulcers in rats.Methods:Fifty male Sprague-Dawley rats aged 6-8 weeks were divided into simple pressure ulcer group and smoking+ pressure ulcer group according to the random number table, with 25 rats in each group. After the rats in the smoking+ pressure ulcer group received passive smoking intervention for 12 weeks, an iron plate was placed in the back muscle of each rat in 2 groups, and a magnet was placed outside the skin at the corresponding position of the iron plate for 2 h at each time, with 5 times a day and continuously for 6 days to reproduce stage 4 pressure ulcer model. Immediately after establishing the model, 3 rats in each group were sacrificed and wound tissue was collected, and hematoxylin-eosin staining was applied to observe the pathological changes of the wounds. On 1, 3, 7, and 14 day (s) after establishing the model, 3 rats in each group were collected to measure the pressure ulcer wound area by the paper jam method. After measurement of the wound area, the rats were sacrificed and the wound tissue was collected, and the protein expression levels of matrix metalloproteinases 9 (MMP-9) and tissue inhibitor of metalloproteinase 1 (TIMP-1) in wound tissue were detected by immunohistochemical method, and the ratio of MMP-9/TIMP-1 was calculated.The wound healing time of the remaining 10 rats in each group was recorded. Data were statistically analyzed with analysis of variance for factorial design, two independent sample t test, and Bonferroni correction. Results:(1) Immediately after establishing the model, muscle fiber necrosis and dissolution with large areas were seen on the wound, the myofibrils arranged loosely, and more lymphocytes and monocytes infiltration were seen around the wound of rats in simple pressure ulcer group. A large number of necrotic myofibers were dissolved and gradually disappeared, the myofibrils arranged loosely, and the number of diffuse lymphocytes and monocyte infiltration in wound of rats in smoking+ pressure ulcer group were significantly higher than those in simple pressure ulcer group. (2) The wound areas of rats in smoking+ pressure ulcer group were significantly larger than those in simple pressure ulcer group on 1, 3, 7, and 14 day (s) after establishing the model ( t=3.019, 2.549, 2.181, 3.674, P<0.05 or P<0.01). (3) On 1 to 14 days after establishing the model, the protein expression levels of MMP-9 and TIMP-1 in the wound tissue and the ratio of MMP-9/TIMP-1 of rats in the two groups increased first and then decreased. On 1, 3, 7, and 14 day (s) after establishing the model, the protein expression levels of MMP-9 in the wound tissue and the ratio of MMP-9/TIMP-1 of rats in smoking+ pressure ulcer group were significantly higher than those in simple pressure ulcer group ( t=4.783, 4.508, 6.325, 7.204, 3.078, 2.989, 4.081, 4.696, P<0.05 or P<0.01), and the protein expression levels of TIMP-1 in wound tissue of rats in the two groups were similar. (4) The wound healing time of rats in smoking+ pressure ulcer group was (48.9±2.6) d, which was significantly longer than (35.2±2.3) d of simple pressure ulcer group ( t=12.477, P<0.05). Conclusions:Smoking can up-regulate the expression of MMP-9 in pressure ulcer wound and result in an imbalance of MMP-9/TIMP-1, thereby affecting the wound healing of stage 4 pressure ulcers in rats.

Objective To investigate the effects of hypoxic preconditioning on learning and memory and the possible protective mechanism in mice with cerebral ischemia-reperfusion injury.Methods Healthy adult male Kunming mice were randomly divided into five groups by Random number table:normal group( N group),hypoxic preconditioning group (HPC group),sham operation group (C group),ischemia-reperfusion group(O group),hypoxic preconditioning and ischemia-reperfusion group(HPC+O group).HPC+O group were given hypoxic preconditioning before 24h of ischemia-reperfusion.The escape latency was detected by Morris water maze and the neuron apoptosis of CA 1 area of hippocampal was determined by immunofluores-cence techniqueR.e sults The escape latency in HPC+O group on the second,third and fourth day of MWM was (39.92±4.52)s,(30.98±2.44)s,(19.69±4.27)s,and significantly lower than that in O group((54.35± 3.66)s,(46.31±4.81)s,(36.81±3.86)s).Mice in HPC+O spent longer time in the target quadrant than that in O group((36.44±5.33)%and(24.5±2.59)%,respectively, P<0.05).Immunofluorescence showed that the apoptotic ration of nerve cells in hippocampal CA 1 was significantly lower than that in O group ( 11.7 ± 0.14 and 1.35±0.14, P<0.05).Conclusion Hypoxic preconditioning can increase hippocampal CA1 neurons hypoxia tolerance of ischemia reperfusion injury in mice,and reduce the incidence of neural cell apoptosis.

AIM: To examine the expression of CD69 on NK cells at the fetomaternal interface in CBA/J?DBA/2 mice as a model of recurrent spontaneous abortion (RSA), and to evaluate the effects of lymphocyte immunotherapy (LIT) on the level of CD69 expression and the relationship with the outcomes of murine fetuses and pups. METHODS: The outcomes of murine fetuses and pups were evaluated in breeding pairs of CBA/J?DBA/2, C57BL/6?DBA/2 and BALB/c?DBA/2 mice. Both preweaning growth curves and Kaplan-Meier survival graphs of pups were constructed throughout postnatal days 1 to 21. In addition, the level of CD69 expression on NK cells at the fetomaternal interface with and without LIT were determined by two-color flow cytometric analysis, stained with PE-CD69 and FITC-DX5. The subpopulation of CD16/CD32 + NK cells was also evaluated. RESULTS: Statistically significant differences were observed between CBA/J?DBA/2 mice and normal fertile controls in the median increase of maternal weight during pregnancy, the number of pups born per litter, the median neonatal weight on postnatal day 1, and the resorption rate of fetuses. The proportion of CD69 +DX5 + cells which represents activated NK cells was significantly higher in CBA/J?DBA/2 mice compared with normal fertile controls, while efficient LIT was able to dramatically decrease the expression of CD69 on NK cells at the fetomaternal interface and this was associated with the decrease of resorption rate accordingly. CONCLUSION: The fraction of CD69 +DX5 + cells seems to be functionally important in the mechanisms by which the embryos were rejected, whereas efficient LIT is capable of reducing the abortion rate via decreasing the expression of CD69 molecules on NK cells at the fetomaternal interface.