Objective:To construct a VP8-mRNA vaccine using human rotavirus spike protein VP8 domain as the immunogen and analyze its immunogenicity in mice.Methods:The VP8-mRNA sequence was designed, optimized, and synthesized. The VP8 gene of rotavirus G1P[8] type was used to construct the plasmid pUC57-VP8-Kan-SapⅠ, which was then sequenced. The plasmid confirmed by sequencing was subjected to large-scale amplification and extraction, followed by linearization, in vitro transcription, and capping. The purified capped products were encapsulated with lipid nanoparticles using a microfluidic control apparatus. The encapsulated VP8-mRNA vaccine was administered intramuscularly to mice at 10, 15, and 20 μg. Serum samples were collected for antibody detection by ELISA. Cellular immune responses were detected by flow cytometry and ELISPOT. Statistical analysis was performed using one-way or two-way analysis of variance and Tukey-Kramer test. Results:The encapsulated VP8-mRNA vaccine was rounded and spherical, with a particle size of about 100 nm, a polymer dispersion index of 0.088, and an encapsulation rate of 92.3%. Two doses of VP8-mRNA vaccine immunization could induce a good immune response in mice. The level of IgG antibody induced after immunization in the 15 μg group was comparable to that of the 20 μg group, and there was no statistical difference ( P>0.05), but the antibody levels in the two groups were significantly higher than that in the 10 μg group ( P<0.000 1). VP8-mRNA vaccine could induce neutralizing antibodies against rotavirus G1 and G9 types. The highest level of neutralizing antibodies against rotavirus type G1 was observed in the 15 μg group, which was significantly higher than that in the 10 μg group ( P<0.05). All immunization groups exhibited good neutralizing ability against rotavirus G9 type. The results of ELISPOT showed that lymphocytes from mice in each vaccine group were able to secrete IFN-γ when stimulated with VP8 peptide. Flow cytometry showed that the proportions of CD8 + T cell subsets in the vaccine groups were higher than that in the control group. Conclusion:The VP8-mRNA vaccine has good immunogenicity in mice and can induce good humoral and T-cell immune responses.

Objective To compare the predictive effectiveness of the Glasgow coma scale(GCS),GCS-pupils scale(GCS-P),Glasgow-Pittsburgh coma scale(GPCS),full outline of unresponsiveness scale(FOUR),and coma recovery scale-revised(CRS-R)in forecasting the prognosis of severe stroke patients.Methods A prospective,consecutive cohort of severe stroke patients admitted to the Department of Neurology,First Medical Center of Chinese PLA General Hospital from September 2021 to April 2024 was enrolled.Demographic and clinical data were collected,including age,sex,length of hospital stay,diagnosis(severe ischemic stroke,severe cerebral hemorrhage,aneurysmal subarachnoid hemorrhage),medical history(hypertension,diabetes,coronary artery disease),smoking and drinking habits,vital signs upon admission(temperature,pulse,respiration,blood pressure),neurological examination findings(including speech and brainstem reflexes)at admission,head imaging results(CT,MRI)within 24 h of admission to assess the presence of brain herniation,and whether intubation occurred within 24 h of admission.Patients underwent GCS,GCS-P,GPCS,FOUR,and CRS-R scoring within 8h of admission.Telephone follow-up was conducted at 6 months post-stroke to assess outcomes using the modified Rankin scale(mRS),with mRS scores of 0-2 classified as the good prognosis group and 3-6 as the poor prognosis group.The receiver operating characteristic(ROC)curve was used to assess the prognostic prediction value of the five scales for poor outcomes at 6 months.The area under the ROC curve(AUC)was calculated,and pairwise comparisons of AUC were performed using the Delong test.Results A total of 179 severe stroke patients were enrolled,including 116 males and 63 females.The group consisted of 132 patients with severe ischemic stroke,30 with severe intracerebral hemorrhage,and 17 with aneurysmal subarachnoid hemorrhage.At 6months,126patients had a poor prognosis and 53 had a good prognosis.(1)There were statistically significant differences in age,temperature,pulse,history of coronary artery disease,smoking and drinking habits,presence of speech impairment,abnormal brainstem reflexes,brain herniation,intubation within 24 h of admission,and GCS,GCS-P,GPCS,FOUR,and CRS-R scores between the poor and good prognosis groups(all P＜0.05).(2)ROC analysis revealed that the AUC(95％CI)for predicting poor outcomes at 6 months in severe stroke patients for GCS,GCS-P,GPCS,FOUR,and CRS-R were 0.808(0.742-0.863),0.815(0.750-0.869),0.828(0.765-0.880),0.841(0.780-0.892),and 0.831(0.768-0.883),respectively.Sensitivities were 76.98％,78.57％,82.54％,84.13％,and 82.54％,and specificities were 73.58％,73.58％,67.92％,71.70％,and 73.58％,respectively.The FOUR had the highest AUC,with an optimal cutoff value of 13.(3)Pairwise comparisons of AUC showed a statistically significant difference between the FOUR and GCS(the difference value of AUC is 0.034,95％CI 0.004-0.064,Z=2.194,P=0.028),but no significant differences were observed between other scales(all P＞0.05).Conclusion Compared to GCS,GCS-P,GPCS,and CRS-R,FOUR may provide more valuable prognostic information for severe stroke patients.

Objective To compare the predictive effectiveness of the Glasgow coma scale(GCS),GCS-pupils scale(GCS-P),Glasgow-Pittsburgh coma scale(GPCS),full outline of unresponsiveness scale(FOUR),and coma recovery scale-revised(CRS-R)in forecasting the prognosis of severe stroke patients.Methods A prospective,consecutive cohort of severe stroke patients admitted to the Department of Neurology,First Medical Center of Chinese PLA General Hospital from September 2021 to April 2024 was enrolled.Demographic and clinical data were collected,including age,sex,length of hospital stay,diagnosis(severe ischemic stroke,severe cerebral hemorrhage,aneurysmal subarachnoid hemorrhage),medical history(hypertension,diabetes,coronary artery disease),smoking and drinking habits,vital signs upon admission(temperature,pulse,respiration,blood pressure),neurological examination findings(including speech and brainstem reflexes)at admission,head imaging results(CT,MRI)within 24 h of admission to assess the presence of brain herniation,and whether intubation occurred within 24 h of admission.Patients underwent GCS,GCS-P,GPCS,FOUR,and CRS-R scoring within 8h of admission.Telephone follow-up was conducted at 6 months post-stroke to assess outcomes using the modified Rankin scale(mRS),with mRS scores of 0-2 classified as the good prognosis group and 3-6 as the poor prognosis group.The receiver operating characteristic(ROC)curve was used to assess the prognostic prediction value of the five scales for poor outcomes at 6 months.The area under the ROC curve(AUC)was calculated,and pairwise comparisons of AUC were performed using the Delong test.Results A total of 179 severe stroke patients were enrolled,including 116 males and 63 females.The group consisted of 132 patients with severe ischemic stroke,30 with severe intracerebral hemorrhage,and 17 with aneurysmal subarachnoid hemorrhage.At 6months,126patients had a poor prognosis and 53 had a good prognosis.(1)There were statistically significant differences in age,temperature,pulse,history of coronary artery disease,smoking and drinking habits,presence of speech impairment,abnormal brainstem reflexes,brain herniation,intubation within 24 h of admission,and GCS,GCS-P,GPCS,FOUR,and CRS-R scores between the poor and good prognosis groups(all P＜0.05).(2)ROC analysis revealed that the AUC(95％CI)for predicting poor outcomes at 6 months in severe stroke patients for GCS,GCS-P,GPCS,FOUR,and CRS-R were 0.808(0.742-0.863),0.815(0.750-0.869),0.828(0.765-0.880),0.841(0.780-0.892),and 0.831(0.768-0.883),respectively.Sensitivities were 76.98％,78.57％,82.54％,84.13％,and 82.54％,and specificities were 73.58％,73.58％,67.92％,71.70％,and 73.58％,respectively.The FOUR had the highest AUC,with an optimal cutoff value of 13.(3)Pairwise comparisons of AUC showed a statistically significant difference between the FOUR and GCS(the difference value of AUC is 0.034,95％CI 0.004-0.064,Z=2.194,P=0.028),but no significant differences were observed between other scales(all P＞0.05).Conclusion Compared to GCS,GCS-P,GPCS,and CRS-R,FOUR may provide more valuable prognostic information for severe stroke patients.

Objective:To construct a VP8-mRNA vaccine using human rotavirus spike protein VP8 domain as the immunogen and analyze its immunogenicity in mice.Methods:The VP8-mRNA sequence was designed, optimized, and synthesized. The VP8 gene of rotavirus G1P[8] type was used to construct the plasmid pUC57-VP8-Kan-SapⅠ, which was then sequenced. The plasmid confirmed by sequencing was subjected to large-scale amplification and extraction, followed by linearization, in vitro transcription, and capping. The purified capped products were encapsulated with lipid nanoparticles using a microfluidic control apparatus. The encapsulated VP8-mRNA vaccine was administered intramuscularly to mice at 10, 15, and 20 μg. Serum samples were collected for antibody detection by ELISA. Cellular immune responses were detected by flow cytometry and ELISPOT. Statistical analysis was performed using one-way or two-way analysis of variance and Tukey-Kramer test. Results:The encapsulated VP8-mRNA vaccine was rounded and spherical, with a particle size of about 100 nm, a polymer dispersion index of 0.088, and an encapsulation rate of 92.3%. Two doses of VP8-mRNA vaccine immunization could induce a good immune response in mice. The level of IgG antibody induced after immunization in the 15 μg group was comparable to that of the 20 μg group, and there was no statistical difference ( P>0.05), but the antibody levels in the two groups were significantly higher than that in the 10 μg group ( P<0.000 1). VP8-mRNA vaccine could induce neutralizing antibodies against rotavirus G1 and G9 types. The highest level of neutralizing antibodies against rotavirus type G1 was observed in the 15 μg group, which was significantly higher than that in the 10 μg group ( P<0.05). All immunization groups exhibited good neutralizing ability against rotavirus G9 type. The results of ELISPOT showed that lymphocytes from mice in each vaccine group were able to secrete IFN-γ when stimulated with VP8 peptide. Flow cytometry showed that the proportions of CD8 + T cell subsets in the vaccine groups were higher than that in the control group. Conclusion:The VP8-mRNA vaccine has good immunogenicity in mice and can induce good humoral and T-cell immune responses.

Objective To observe the value of 68Ga-DOTA-NOC PET/CT for diagnosing pheochromocytoma(PCC)and paraganglioma(PGL).Methods Thirty-eight patients with suspected or confirmed PCC/PGL who underwent 68 Ga-DOTA-NOC PET/CT were retrospectively enrolled,among them 20 cases underwent 131I-metaiodobenzylguanidine(MIBG)SPECT/CT during the same period.The value of 68Ga-DOTA-NOC PET/CT for diagnosing PCC/PGL at individual and lesion levels were analyzed and compared to the results of 131I-MIBG SPECT/CT.Results Among 38 cases,there were 20 cases of PCC,14 cases of PGL,1 case of adrenocortical carcinoma and 3 cases of benign adrenal hyperplasia.The sensitivity,specificity,positive predictive value,negative predictive value and accuracy of 68 Ga-DOTA-NOC PET/CT for diagnosing PCC/PGL in all 38 cases was 87.88％(29/33),60.00％(3/5),93.55％(29/31),42.86％(3/7)and 84.21％(32/38),respectively.Totally 188 lesions were detected in 34 cases,with detection rate of 89.95％(188/209).For 20 patients who underwent both 2 kinds examinations,the detection rate of bone,lymph node,liver,lung metastases and the overall lesions of 68Ga-DOTA-NOC PET/CT were all higher than those of 131I-MIBG SPECT/CT(all P＜0.05).No significant difference of diagnostic accuracy of PCC/PGL was found between 68 Ga-DOTA-NOC PET/CT and 131I-MIBG SPECT/CT(P＞0.05).Conclusion The value of 68 Ga-DOTA-NOC PET/CT for diagnosing PCC/PGL was comparable to that of 131I-MIBG SPECT/CT,but the former showed higher detection rate of metastases,hence being helpful to staging and risk stratification of PCC/PGL.

Objective To observe the value of 68Ga-DOTA-NOC PET/CT for diagnosing pheochromocytoma(PCC)and paraganglioma(PGL).Methods Thirty-eight patients with suspected or confirmed PCC/PGL who underwent 68 Ga-DOTA-NOC PET/CT were retrospectively enrolled,among them 20 cases underwent 131I-metaiodobenzylguanidine(MIBG)SPECT/CT during the same period.The value of 68Ga-DOTA-NOC PET/CT for diagnosing PCC/PGL at individual and lesion levels were analyzed and compared to the results of 131I-MIBG SPECT/CT.Results Among 38 cases,there were 20 cases of PCC,14 cases of PGL,1 case of adrenocortical carcinoma and 3 cases of benign adrenal hyperplasia.The sensitivity,specificity,positive predictive value,negative predictive value and accuracy of 68 Ga-DOTA-NOC PET/CT for diagnosing PCC/PGL in all 38 cases was 87.88％(29/33),60.00％(3/5),93.55％(29/31),42.86％(3/7)and 84.21％(32/38),respectively.Totally 188 lesions were detected in 34 cases,with detection rate of 89.95％(188/209).For 20 patients who underwent both 2 kinds examinations,the detection rate of bone,lymph node,liver,lung metastases and the overall lesions of 68Ga-DOTA-NOC PET/CT were all higher than those of 131I-MIBG SPECT/CT(all P＜0.05).No significant difference of diagnostic accuracy of PCC/PGL was found between 68 Ga-DOTA-NOC PET/CT and 131I-MIBG SPECT/CT(P＞0.05).Conclusion The value of 68 Ga-DOTA-NOC PET/CT for diagnosing PCC/PGL was comparable to that of 131I-MIBG SPECT/CT,but the former showed higher detection rate of metastases,hence being helpful to staging and risk stratification of PCC/PGL.

Objective To investigate the clinical data,brain pathology and molecular genetic characteristics of retinal vascular disease families with leukoencephalopathy and systemic damage.Methods The clinical data of two families of RVCL-S(retinal vasculopathy with cerebral leukoencephalopathy and systemic manifestations,RVCL-S)were collected and family maps were drawn to analyze the clinical manifestations,imaging,brain pathology and molecular genetic characteristics.Results Family 1:there were 3 male cases and the age of onset was 10 years,29 years and 40 years,respectively.Family 2:there was one male case and the age of onset was 32 years.Both families presented with retinal vascular disease,leukoencephalopathy and multi-system damage,the latter including liver,kidney and digestive tract involvement.There were 4 asymptomatic carriers in two families.The cranial CT of family 1-Ⅱ2 showed lamellar low density near the posterior corner of the left ventricle with multiple intracranial high density calcification.Brain MRI plain scan showed lateral ventricle lesions.The brain MRI plain and enhancement scans of family 1-I5 showed frontotemporal cortex lesions with peripheral edema and space occupying effect,and the ring enhancement was remarkable.The brain MRI plain scan and enhancement scan of family 2-Ⅱ1 showed the right and left frontal lobe lesions,accompanied by peripheral edema and enhancement,and the occupying effect was obvious.The operative pathology of brain tissue from family 1-I5 showed endothelial cell hyperplasia.The pathological manifestations of family 2-Ⅱ1 encephalopathy were consistent with"cerebral infarction"after two operations.The small blood vessels in the small intestinal wall showed endothelial cell proliferation.Molecular genetics:TREX1 D272Rfs heterozygous mutation was present in family 1-Ⅱ2,and his offspring including two daughters and one son were asymptomatic mutation carriers.TREX1 S246Ifs heterozygous mutation was detected in the 2-Ⅱ1 gene of the family which was not found in either the father or the mother found the mutation,and the son was asymptomatic carrier of the mutation.Conclusion The main clinical manifestations of RVCL-S are retinal vascular disease,nervous system involvement and multi-system damage.Imaging findings showed that intracranial lesions were space-occupying,accompanied by peripheral edema and enhancement.The pathological features were small vessel endothelial cell proliferation and lumen stenosis.Genetic results confirmed the presence of TREX1 gene mutation.

Objective:To investigate the clinical application of 68Ga-cyclo( L-arginylglycyl- L-α-aspartyl- D-tyrosyl-N6-(((4, 7-bis(carboxymethyl)-1, 4, 7-triazonan-1-yl)acetyl))- L-lysyl) (NODAGA-RGD) PET/CT to evaluate short-term efficacy of tyrosine kinase inhibitor (TKI) in distant metastatic differentiated thyroid cancer (dmDTC). Methods:From October 2019 to March 2023, 13 dmDTC patients (5 males, 8 females; age: 68(65, 69) years) from Nanjing First Hospital were retrospectively enrolled, of which 9 were clinically confirmed as radioactive iodine-refractory differentiated thyroid cancer (RAIR-DTC) and 4 were dmDTC without radioactive iodine treatment. All patients underwent 68Ga-NODAGA-RGD PET/CT to assess neovascularization of the target lesions (TL), and the SUV max and target background ratio (T/B) were recorded. After 3 months of TKI treatment (anrotinib ( n=9) or apatinib ( n=4)), change rates of the maximum diameter of TL and thyroglobulin (Tg) were measured. The correlation of SUV max, T/B and the change rate of the maximum diameter of TL were analyzed by Spearman rank correlation analysis. ROC curve analysis was performed for the effectiveness of the T/B and TKI therapy, and the difference of the remission rate of lesions was analyzed by Fisher exact test. Results:In 13 patients, 36 TL were measured by 68Ga-NODAGA-RGD PET/CT with SUV max of 5.44(3.43, 7.56) and T/B of 5.25(4.50, 7.23). The change rate of the maximum diameter of TL was -30%(-39%, -21%) and the change rate of Tg was -68%(-96%, -52%). T/B was negatively correlated with the change rate of the maximum diameter of TL after TKI therapy ( rs=-0.46, P=0.005), while SUV max was not correlated with the change rate of the maximum diameter of TL ( rs=0.03, P=0.883). ROC curve analysis showed that the optimal cut-off value for T/B was 4.95, with the AUC of 0.698, the sensitivity of 87.5%, and the specificity of 60.0%. Compared to lesions with T/B<4.95, those with T/B≥4.95 showed higher remission rate (2/14 vs 63.6%(14/22); P=0.006). After 3 months of TKI treatment, the disease control rate was 12/13. Conclusion:68Ga-NODAGA-RGD PET/CT can effectively reflect tumor neovascularization, predict efficacy of TKI therapy, and provide powerful imaging evidence for TKI therapy in dmDTC.

To report a typical case of Morvan syndrome with positive anti-leucine rich glioma-inactivated 1(LGI1) and contactin-associated protein 2 (CASPR2) antibodies in serum and cerebrospinal fluid. A 39-years-old female initially presented weakness of extremeties. The main symptoms included paroxysmal limb pain, wheezing, itching, muscle twitching, epilepsy, hypomnesia, dysphoria, apathy, intractable insomnia, salivation and sweating. Tests of electrolytes found hypokalemia (2.7-3.1 mmol/L) and hyponatremia (130-136 mmol/L). Arterial blood gas analysis showed hypoxemia (oxygen saturation 50%-70%). Total thyroxine (TT4) was elevated to 207 nmol/L with positive thyroid peroxidase antibody (TPO-Ab) and thyroglobulin antibody (TG-Ab). LGI1and CASPR2 antibodies (CBA method) were positive in both serum and cerebrospinal fluid, and the remaining antibodies related to autoimmune encephalitis and paraneoplastic syndrome were negative. Head MRI was almost normal, while mild abnormalities were found in electroencephalogram. Electromyography showed slightly increased voltage of left quadriceps motor unit potential. After treated with corticosteroids, IVIG and mycophenolate mofetil, the patient completely improved. Cognitive function scores recovered from MoCA/MMSE (16/24) to MoCA/MMSE (26/29). Positivity of LGI1/CASPR2 antibodies both in serum/cerebrospinal fluid are rarely seen in patients with Morvan syndrome. Steroids and immunosuppressants are suggested for treatment as early as possible.

Objective:To explore the normal ranges of perfusion parameters between cerebral hemisphere, cerebellar hemisphere and brain anatomical subregions (56 pairs) in different gender and age groups with multiple post labeling delay time (Multi-PLD) arterial spin labeling (ASL) imaging.Methods:From November 2020 to December 2020, 42 healthy adult volunteers (Male 25, Female 17) were recruited to perform 7 PLD ASL imaging, including 21 young adults (15 males and 6 females, aged 23—35 years) and 21 seniors (10 males and 11 females, aged 36—74 years). The data was processed offline by Cereflow software to obtain arterial arrival time (ATT) and corrected cerebral blood flow (CBF) and cerebral blood volume (CBV) perfusion parameters. SimpleITK standardization function was used to standardize the calculated perfusion image according to the anatomical automatic labeling (AAL) template. Therefore, CBF, ATT, CBV perfusion values of brain subregions were obtained. Paired samples t test, Wilcoxon rank sum test, independent samples t test and Mann-Whitney U test were used to compare the differences of perfusion parameters in the cerebral hemisphere, the cerebellar hemisphere, brain subregions depending on side, gender and age. Pearson correlation analysis was used to compare the correlations of perfusion parameters with age. Results:CBF in 62.5% (35/56) subregions and CBV in 44.6% (25/56) subregions were higher in right side than those in left side. ATT in most brain anatomical subregions (16/56) were higher in left side. The CBF [(35.30±8.31) vs. (34.34±7.53) ml·100g -1·min -1, P=0.021], CBV [(0.47±0.11) vs. (0.45±0.09) ml/100g, P<0.001], ATT [(1.30±0.10) vs. (1.24±0.11) s, P<0.001] in left cerebellar hemisphere were higher than that of right side. The CBF (28/56) of cerebral hemisphere, cerebellar hemisphere and brain subregions was higher in females than that in males, while ATT in 83.9% (47/56) subregions was lower than that in males (all P<0.05). CBV in female subjects was higher only in 5 brain regions (superior occipital gyrus, middle occipital gyrus, inferior occipital gyrus, superior parietal gyrus and cerebelum_7b) (all P<0.05). In young subjects, CBF in 44.6% (25/56) subregions and CBV in 33.9% (19/56) subregions were higher than those in the senior group (all P<0.05). The ATT in most subregions in young group were lower than those in senior group, but the difference was statistically significant only in rectus gyrus ( P=0.026) and paracentral lobule ( P=0.006). The CBF ( r=-0.430, P=0.005) and CBV ( r=-0.327, P=0.035) of cerebral hemisphere were negatively correlated with age. The CBF (24/25, r range:-0.497 —-0.343, all P<0.05) and CBV (16/19, r range:-0.474 —-0.322, all P<0.05) in most subregions were negatively correlated with age, while ATT was positively correlated (gyrus rectus: r=0.311, P=0.045; paracentral lobule: r=0.392, P=0.010). Conclusions:Multi-PLD ASL imaging could be applied for quantitative analysis of brain perfusion. The perfusion parameters of anatomical subregions are different depending on side, gender, and age.