OBJECTIVE To investigate the mechanism by which Naozhenning granules （NZN） improve learning and memory impairment in a rat model of multiple cerebral concussion （MCC）. METHODS The MCC rat model was established using the closed controlled cortical impact method. The experiment was set up with a blank group （normal saline）， a model group （normal saline）， a piracetam group （positive control group， 0.324 g/kg）， and high-， medium-， and low-dose NZN groups （5.4， 2.7， 1.35 g/kg）， with 11 rats in each group. Drugs or normal saline were administered by gavage once daily for 28 consecutive days. General condition and body weight were monitored throughout the experiment. The sucrose preference rate and novel object recognition index were measured； Evans blue （EB） extravasation in the cerebral cortex was detected； pathological changes of cortical neurons were observed； the levels of B-cell lymphoma-2 （Bcl-2）， Bcl-2-associated X protein （Bax）， interleukin-6 （IL-6）， IL-10， and tumor necrosis factor-α （TNF-α） in the cerebral cortex were determined； and the phosphorylation levels of AMP-activated protein kinase （AMPK）， glycogen synthase kinase 3β （GSK3β）， and Tau protein were detected. RESULTS Compared with the blank group， the model group showed poor mental state， sluggish response to external stimuli， reduced food and water intake， decreased limb flexibility， and disheveled fur. Body weight， sucrose preference rate， and novel object recognition index were significantly decreased （ P ＜0.05）； EB extravasation in the cerebral cortex was significantly increased （ P ＜0.05）， with severe neuronal damage. The positive area ratio of Bax protein， IL-6 and TNF-α levels， and Tau protein phosphorylation level were all significantly increased （ P ＜0.05）， whereas the positive area ratio of Bcl-2 protein， IL-10 level， and AMPK and GSK3β protein phosphorylation levels were significantly decreased （ P ＜0.05）. Compared with the model group， all NZN dose groups showed improvements in general condition and pathological damage， with quantitative indices partially restored， and the differences in quantitative indices in high-dose NZN group were statistically significant （ P ＜0.05）. CONCLUSIONS NZN can effectively improve learning and memory impairment in MCC model rats. The mechanism may be related to activating the AMPK/GSK3β pathway， inhibiting inflammatory response， reducing Tau protein phosphorylation level， and then repairing the neuronal injury.

Objective: To investigate the prevalence of Dengue virus (DENV) infection among voluntary blood donors in Xishuangbanna Dai Autonomous Prefecture, and to evaluate the necessity of implementing nucleic acid testing (NAT) for blood donors during the rainy season (May-October). Methods: Prior to initiating donor screening, the Xishuangbanna Central Blood Center conducted in-house validation of reagent performance and participated in external quality assessment (EQA) organized by the National Center for Clinical Laboratories (NCCL). During the surveillance period (August-October 2024), a total of 2 919 donor samples were screened using a 6-sample mini-pool NAT strategy. Daily internal quality controls were recorded. Samples that tested positive in pooled screening were deconvoluted and retested in duplicate; only those reactive in both replicate wells were sent to the NCCL for confirmatory testing. At NCCL, samples underwent re-testing using five domestic NAT reagents, as well as serological assays for NS1 antigen and DENV-specific IgG/IgM. Confirmed positive samples were further characterized by serotyping, envelope (E) gene sequencing, and phylogenetic analysis using the maximum likelihood method. Results: The DENV NAT reagent demonstrated consistent detection of 40 copies/mL controls in individual donor (ID)-NAT test (mean CT: 35.61±0.40). During the 63-day quality control monitoring, DENV detection remained stable (mean CT: 22.53±0.72). The center achieved full marks in EQA assessments for 2023 and 2024. Three reactive pools were identified in initial screening, and subsequent individual testing confirmed three DENV RNA-positive donors (sample numbers: 2401, 2402, and 2403). The confirmatory test results from NCCL were: all five NAT platforms consistently detected DENV RNA in the three samples; for serological tests, 2 samples (2402, 2403) were positive for NS1 antigen, while all three samples were negative for both IgG and IgM antibodies. DENV serotyping reagents identified DENV-2 in all cases, which were further confirmed as DENV-2 Genotype Ⅱ-Cosmopolitan by E gene sequencing. Phylogenetic analysis indicated that samples 2401 and 2402 clustered with Southeast Asian strains (Thailand/MZ636802.1, Laos/PQ775621.1), while sample 2403 closely matched a previously reported local Yunnan strain (PV544686.1). Conclusion: DENV-2 infection was detected among blood donors in Xishuangbanna during the rainy season, indicating concurrent risks of imported and local transmission. We recommend implementing pooled NAT screening for blood donors in high-risk areas during dengue epidemic seasons, along with strengthened laboratory quality control, to enhance blood safety.

Objective To screen a Madin-Darby canine kidney (MDCK) cell line for H5N1 influ-enza virus isolation and to evaluate its safety in vaccine production. Methods MDCK cells were cloned by the method of limiting dilution. Hemagglutination test was used to screen MDCK cells that were suitable for H5N1 influenza virus production. Tests for analyzing the characteristics, extraneous agents, endogenous agents and tumorigenicity of MDCK cells were performed according to Chinese Pharmacopeia Volume Ⅲ. Results A total of 108 MDCK cell lines were obtained and three of them were selected after hemagglutina-tion test. G1 cells were chosen following further screening with tumorigenicity test and receptor abundance analysis. The average number of chromosomes of the MDCK-G1 cells was 78±4. No bacteria, fungi or myco-plasma contamination was detected. In experimental group, each nude mouse was injected with 1×107/ml viable cells to observe their tumorigenicity. Twelve weeks after cell injection, no node was found at injection sites or in gross anatomy. There was no significant difference between the experimental and negative control groups. The result of the tumorigenicity test was negative. No node formation was found after injecting nude mice with cell lysate or cellular DNA collected from equivalent amount of cells. It was indicated that the MDCK-G1 cells were of low-oncogenic potential. Conclusions The MDCK-G1 cell line could be used as a substrate to produce H5N1 influenza virus vaccine.

Objective To investigate the characteristics of cognitive impairment and psychotic symptoms in general paresis of insane (GPI) before and after penicillin therapy, and explore factors that may predict the clinical outcomes. Methods Thirty patients with GPI were recruited. All GPI patients underwent a comprehensive neuropsychological assessment before receiving penicillin therapy, and returned for follow-up visits after 7 months. The severity of dementia was determined by Clinical Dementia Rating Scale (CDR), cognitive functions were assessed by Mini Mental State Examination (MMSE) and Alzheimer 's Disease Assessment Scale-cognitive subscale (ADAS-Cog), ability of daily living was assessed by Instrumental Activities of Daily Living Scale (IALD) and Physical Self maintenance Scale(PSMS), behavioral and psychological symptoms were assessed by Neuropsychiatric Inventory (NPI). Aqueous crystalline penicillin G 24 million units per day was administered as continuous infusion for 14 days, followed by benzathine penicillin 2.4 million units IM once per week for 3 weeks. Patients returned for follow-up visits after 7 months. Clinical outcomes were determined by the improvement of neuropsychological test scores at the end of the treatment. Grouped by CDR scores, changes in neuropsychological tests scores among different GPI groups were used to explore the correlation between severity of dementia and clinical outcomes. Univariate analysis and multivariate linear regression analysis were used to identify factors that may predict the clinical outcomes. Results (1)After penicillin therapy, GPI patients' MMSE scores(14.4± 6.9 vs.17.1 ± 9.1)and IADL scores(4.0(2.0, 5.0)vs.6.0(2.0, 7.3))both improved significantly(t=5.820, Z=3.710, P<0.01),while in ADAS-Cog, only factor scores of attention(1.5(0.7, 3.0)vs.1.5(0, 2.3))reduced significantly(Z=- 2.680, P<0.01). NPI's total scores(46.0 ± 27.1 vs.17.6 ± 15.4)and subscores of hallucination, delusion, agitation, depression, euphoria, disinhibition and irritability reduced significantly (Z=-4.940,-2.381,-2.504,-3.095,-2.492,-3.097,-2.527,-3.715, all P<0.05).(2) Grouped by the CDR scores, MMSE scores and IADL scores in very mild GPI group with CDR=0.5 improved significantly. In mild GPI group with CDR=1, significant changes were also found in all neuropsychological tests scores(MMSE,t=5.409, P<0.01), total scores of ADAS-Cog (Z=-2.366,P<0.05), IADL (Z=2.546, P<0.05), total scores of NPI (Z=-3.558,P<0.01), but except for PSMS. In moderate to severe GPI group with CDR>1,significant change was found only in total scores of NPI (t=-3.772,P<0.05). (3) Univariate analysis and multivariate linear regression analysis showed that improvement of MMSE scores after the treatment was significantly correlated with IADL scores and MMSE scores at baseline(β=0.541,P=0.004;β=0.364,P=0.044). Conclusions After penicillin treatment, GPI patients may improve in both cognitive function and psychotic symptoms but not in all the domains. Symptoms of anxiety, sleep/nigh-time behavior change, and apathy, as well as moderate to severe GPI patients may not benefit much from the treatment.

Objective To investigate the characteristics of cognitive impairment and psychotic symptoms in general paresis of insane (GPI) before and after penicillin therapy, and explore factors that may predict the clinical outcomes. Methods Thirty patients with GPI were recruited. All GPI patients underwent a comprehensive neuropsychological assessment before receiving penicillin therapy, and returned for follow-up visits after 7 months. The severity of dementia was determined by Clinical Dementia Rating Scale (CDR), cognitive functions were assessed by Mini Mental State Examination (MMSE) and Alzheimer 's Disease Assessment Scale-cognitive subscale (ADAS-Cog), ability of daily living was assessed by Instrumental Activities of Daily Living Scale (IALD) and Physical Self maintenance Scale(PSMS), behavioral and psychological symptoms were assessed by Neuropsychiatric Inventory (NPI). Aqueous crystalline penicillin G 24 million units per day was administered as continuous infusion for 14 days, followed by benzathine penicillin 2.4 million units IM once per week for 3 weeks. Patients returned for follow-up visits after 7 months. Clinical outcomes were determined by the improvement of neuropsychological test scores at the end of the treatment. Grouped by CDR scores, changes in neuropsychological tests scores among different GPI groups were used to explore the correlation between severity of dementia and clinical outcomes. Univariate analysis and multivariate linear regression analysis were used to identify factors that may predict the clinical outcomes. Results (1)After penicillin therapy, GPI patients' MMSE scores(14.4± 6.9 vs.17.1 ± 9.1)and IADL scores(4.0(2.0, 5.0)vs.6.0(2.0, 7.3))both improved significantly(t=5.820, Z=3.710, P<0.01),while in ADAS-Cog, only factor scores of attention(1.5(0.7, 3.0)vs.1.5(0, 2.3))reduced significantly(Z=- 2.680, P<0.01). NPI's total scores(46.0 ± 27.1 vs.17.6 ± 15.4)and subscores of hallucination, delusion, agitation, depression, euphoria, disinhibition and irritability reduced significantly (Z=-4.940,-2.381,-2.504,-3.095,-2.492,-3.097,-2.527,-3.715, all P<0.05).(2) Grouped by the CDR scores, MMSE scores and IADL scores in very mild GPI group with CDR=0.5 improved significantly. In mild GPI group with CDR=1, significant changes were also found in all neuropsychological tests scores(MMSE,t=5.409, P<0.01), total scores of ADAS-Cog (Z=-2.366,P<0.05), IADL (Z=2.546, P<0.05), total scores of NPI (Z=-3.558,P<0.01), but except for PSMS. In moderate to severe GPI group with CDR>1,significant change was found only in total scores of NPI (t=-3.772,P<0.05). (3) Univariate analysis and multivariate linear regression analysis showed that improvement of MMSE scores after the treatment was significantly correlated with IADL scores and MMSE scores at baseline(β=0.541,P=0.004;β=0.364,P=0.044). Conclusions After penicillin treatment, GPI patients may improve in both cognitive function and psychotic symptoms but not in all the domains. Symptoms of anxiety, sleep/nigh-time behavior change, and apathy, as well as moderate to severe GPI patients may not benefit much from the treatment.

Objective To explore the plasma levels of soluble CD40 (sCD40) and soluble CD40 ligand (sCD40L) in the patients with Alzheimer’s disease (AD) and those with amnestic mild cognitive impairment (aMCI). Methods The levels of plasma sCD40 and sCD40L were measured in 20 patients with AD, 35 patients with aMCI, and 32 cognitively normal controls (NC) using commercially available ELISAs. The cognitive function of AD and aMCI patients was mea?sured by mini-mental state examination (MMSE). Results There were significant differences in plasma sCD40 among AD, aMCI and NC groups (P<0.05) as the medians (the upper and lower quartiles) of plasma levels were 123.3 (97.4, 149.5) pg/mL, 102.9 (63.6, 124.0) pg/mL and 70.66 (51.0, 90.8) pg/mL, respectively. There were significant differences in plasma sCD40L among AD, aMCI and NC groups (P<0.05) as plasma levels were 537.0 (316.0, 1134.0) pg/mL, 316.0 (190.0,546.0) pg/mL and 167.0 (107.5,478.0) pg/mL. A negative correlation between the plasma concentrations of sCD40L and the MMSE scores was found in aMCI patients (r=-0.736, P<0.001). Conclusions There are relevant chang?es of plasma sCD40 and sCD40L levels in patients with AD and aMCI. The present results suggest that plasma levels of sCD40 and sCD40L may be appropriate biomarkers for AD patients and indicate that CD40-CD40L signaling may be in?volved in AD pathophysiology.

Objective We aimed to use 1H-MRS to characterize metabolite concentrations in the bilateral hippo?campus in GPI patient. Methods Metabolite ratios in the bilateral hippocampus were compared between patients with GPI (n=52) and Normal Control (NC) (n=38). Clinical neurological tests were measured in all subjects and were correlat?ed to the metabolite concentrations. Results The GPI patients showed significantly lower concentrations of N-acetylas?partate(NAA)/creatine(Cr) ratios in the bilateral hippocampus region compared to the NC subjects. There was significant?ly difference in the NAA/Cr ratios between the mild GPI dementia and the severe GPI dementia groups on the left hippo?campus region. We found that the NAA/Cr ratios concentrations were positively correlated with Mini Mental state Exami?nation (MMSE) and Montreal Cognitive Assessment scale (MoCA) scores in the left hippocampus region and the mI/Cr ra?tios concentrations were positively correlated. Conclusions GPI Patients have neuronal dysfunction in the bilateral hippo?campus. The severity of cognitive impairments is associated with the severity of the damage in the left side.