OBJECTIVES: To investigate the effects of early life intervention with Bifidobacterium bifidum BD-1 (B. bifidum BD-1) on hyperactivity in a female mouse model of attention deficit hyperactivity disorder (ADHD) and explore the underlying mechanisms. METHODS: Eight newborn female Wistar-Kyoto (WKY) rats and 6 spontaneous hypertensive rats (SHRs) were gavaged with saline and another 6 SHRs were gavaged with B. bifidum BD-1 (10⁹ CFU) daily for 3 weeks. Open field test of the rats was conducted at 7 weeks, and fecal samples were collected at weaning (3 weeks) and at 7 weeks for 16S rRNA sequencing. Immunofluorescent staining was used to detect dopamine transporter (DAT) and tyrosine hydroxylase (Th) levels in the striatum and activated microglia in the prefrontal cortex. Treg cells in the mesenteric lymph nodes, spleen and blood were analyzed using flow cytometry. RESULTS: The SHRs traveled a significantly greater distance in open fields test than WKY rats, and this behavior was significantly attenuated by B. bifidum BD-1 intervention. The expression of DAT and Th in the striatum was significantly lower in the SHRs than in WKY rats, while B. bifidum BD-1 treatment obviously increased Th levels in the SHRs. B. bifidum BD-1 intervention significantly deceased the number of activated microglia and increased Treg cell counts in the spleen of SHRs. The treatment also enhanced α diversity in gut microbiota of the SHRs and resulted in a decreased Firmicutes/Bacteroidota ratio, more active Muribaculaceae growth, and suppression of Clostridia_UCG-014 proliferation. CONCLUSIONS Early life intervention with B. bifidum BD-1 alleviates hyperactivity in female SHRs by modulating the gut microbiota and peripheral immune response, suppressing neuroinflammation and improving dopaminergic system function. These findings provide evidence for early prevention strategies and support the development and application of psychobiotics for ADHD.
Animals ; Female ; Rats ; Rats, Inbred WKY ; Rats, Inbred SHR ; Attention Deficit Disorder with Hyperactivity/therapy* ; Bifidobacterium bifidum ; Probiotics/therapeutic use* ; Dopamine Plasma Membrane Transport Proteins/metabolism* ; Tyrosine 3-Monooxygenase/metabolism* ; Gastrointestinal Microbiome ; Disease Models, Animal

Objective To make a preliminary investigation of the effect of the immune pathway mediated by live Lacticaseibacillus paracasei N11 15 on the development of primary hippocampal neurons cultured in vitro.Methods Live Lacticaseibacillus paracasei N1115 suspension of an appropriate concentration was used as the experimental group.Peptidoglycan(PGN)and lipopolysaccharide(LPS)were used as positive controls,and RPMI1640 medium served as the blank control.These were co-cultured with RAW264.7 cells to obtain the co-culture mediums and the total cellular RNA,and to measure the expression and secretion of cytokines.After centrifugation,the supernatants were co-cultured with primary hippocampal neurons at appropriate ratios.The co-culture mediums were collected,and the total cellular RNA was extracted to measure the expression of genes related to synaptic development in neurons.Following immunofluorescence staining of the primary hippocampal neurons,the presynaptic and presynaptic membrane-associated proteins,including synaptophysin(SYP)and the postsynaptic density protein 95(PSD95),and neuronal cell maturation markers,including microtubule-associated protein 2(MAP-2),and doublecortin(DCX)were quantitatively analyzed.Additionally,the morphological development of the neurons were measured.Results Compared with the blank control,the mRNA expression levels of interleukin(IL)-6 and tumor necrosis factor-α(TNF-α)increased by 7471％and 926％,respectively,after the RAW264.7 cells were treated with live Lacticaseibacillus paracasei N1115,while their secretion levels increased by 184.16 pg/mL and 12320.76 pg/mL,respectively,all showing statistically significant differences(P＜0.05).The activation ability of N1115 live bacteria was stronger than that of PGN but weaker than that of LPS,showing statistically significant differences(P＜0.05).Compared with the blank control,following the intervention with the supernatant from the co-culture of N1115 live bacteria and RAW264.7 cells,the viability of primary hippocampal neurons in the 10％supernatant intervention group increased by 19.25％,showing statistically significant differences(P＜0.05),and the mRNA expression of SYP and PSD95 increased by 137％and 159％,respectively,showing statistically significant differences(P＜0.05).The total neurite length(489.88 μm)of the neurons of the group intervened with the supernatant of N11 15 live bacteria was increased compared to that of the blank control group(381.51 μm),and the cell body area(2092.22 μm2),maximum neurite length(184.78 μm),total neurite length,average neurite length(108.38 μm),and branching points(4.84 s)were higher than those in the two positive control groups,showing statistically significant differences(P＜0.05).Conclusion Lacticaseibacillus paracasei N1115 can significantly activate the immune regulatory function of macrophages,thereby promoting the morphological development and synaptic function of nerve cells.

Objective:To investigate the correlation between the age of men and the quality of semen, as well as the DNA fragmentation index (DFI) in assisted reproduction couples.Methods:A cross-sectional study was conducted, collecting clinical data from 3 361 men who underwent assisted reproductive technology for fertility treatment at the Reproductive Medicine Center of Wuhan University People's Hospital from January 2020 to August 2023 due to non-male factors. Computer-assisted semen analysis system (CASA) was used to analyze semen quality parameters, and DFI was measured by Sperm Chromatin Structure Assay (SCSA). Patients were categorized into three groups based on age: 20-29 years (group A), 30-39 years (group B), and 40-59 years (group C). Multiple linear regression and restricted cubic spline (RCS) were used to analyze the correlation between age and semen quality parameters as well as DFI.Results:The study included 3 361 participants with an average age of (34.86±5.34) years. In groups B and C, sperm progressive motility [(40.17±18.16)%, (37.83±16.96)%] and total motility [(55.27±21.37)%, (53.09±21.14)%] were significantly lower than those in group A [(43.78±18.16)%, (58.29±20.24)%], sperm concentration [(78.96±63.04)×10 6/mL, (91.93±72.28)×10 6/mL] and DFI [(19.18±12.18)%, (21.73±12.52)%] were significantly higher than those in group A [(71.75±57.44)×10 6/mL, (16.31±10.04)%], semen volume was significantly lower in group C [(2.94±1.42) mL] than in groups B and A [(3.28±1.43) mL, (3.15±1.58) mL], with statistically significant differences (all P<0.05). After adjusting for education level, body mass index, smoking, alcohol consumption, and abstinence days, the results of multiple linear regression showed that compared with group A, the age of group B was negatively associated with sperm progressive motility ( β=-3.055, 95% CI: -4.879--1.231) and total motility ( β=-2.366, 95% CI:-4.516--0.216), while positively associated with sperm concentration ( β=7.752, 95% CI: 1.398-4.106) and DFI ( β=2.744, 95% CI: 1.526-3.961). Compared with group A, the age of group C was negatively correlated with semen volume ( β=-0.379, 95% CI: -0.565--0.192), sperm progressive motility ( β=-5.507, 95% CI: -7.714--3.301), and total motility ( β=-4.932, 95% CI: -7.532--2.331), while positively correlated with sperm concentration ( β=17.288, 95% CI: 9.604-24.973) and DFI ( β=5.226, 95% CI: 3.753-6.699). The results of RCS analysis indicated a significant linear dose-response relationship between age and semen volume ( Pnon-linearity=0.424), showing a decline in semen volume with increasing age ( P<0.001). Non-linear dose-response relationships were observed between age and sperm density ( Pnon-linearity=0.003), sperm progressive motility ( Pnon-linearity<0.001), total motility ( Pnon-linearity<0.001), and DFI ( Pnon-linearity=0.004). Conclusion:In men undergoing assisted reproductive technology for fertility treatment due to non-male factors, age greater than 30 years is significantly associated with a decrease in semen volume, sperm progressive motility, and total motility, as well as an increase in sperm density and DFI.

Objective:To investigate the correlation between the age of men and the quality of semen, as well as the DNA fragmentation index (DFI) in assisted reproduction couples.Methods:A cross-sectional study was conducted, collecting clinical data from 3 361 men who underwent assisted reproductive technology for fertility treatment at the Reproductive Medicine Center of Wuhan University People's Hospital from January 2020 to August 2023 due to non-male factors. Computer-assisted semen analysis system (CASA) was used to analyze semen quality parameters, and DFI was measured by Sperm Chromatin Structure Assay (SCSA). Patients were categorized into three groups based on age: 20-29 years (group A), 30-39 years (group B), and 40-59 years (group C). Multiple linear regression and restricted cubic spline (RCS) were used to analyze the correlation between age and semen quality parameters as well as DFI.Results:The study included 3 361 participants with an average age of (34.86±5.34) years. In groups B and C, sperm progressive motility [(40.17±18.16)%, (37.83±16.96)%] and total motility [(55.27±21.37)%, (53.09±21.14)%] were significantly lower than those in group A [(43.78±18.16)%, (58.29±20.24)%], sperm concentration [(78.96±63.04)×10 6/mL, (91.93±72.28)×10 6/mL] and DFI [(19.18±12.18)%, (21.73±12.52)%] were significantly higher than those in group A [(71.75±57.44)×10 6/mL, (16.31±10.04)%], semen volume was significantly lower in group C [(2.94±1.42) mL] than in groups B and A [(3.28±1.43) mL, (3.15±1.58) mL], with statistically significant differences (all P<0.05). After adjusting for education level, body mass index, smoking, alcohol consumption, and abstinence days, the results of multiple linear regression showed that compared with group A, the age of group B was negatively associated with sperm progressive motility ( β=-3.055, 95% CI: -4.879--1.231) and total motility ( β=-2.366, 95% CI:-4.516--0.216), while positively associated with sperm concentration ( β=7.752, 95% CI: 1.398-4.106) and DFI ( β=2.744, 95% CI: 1.526-3.961). Compared with group A, the age of group C was negatively correlated with semen volume ( β=-0.379, 95% CI: -0.565--0.192), sperm progressive motility ( β=-5.507, 95% CI: -7.714--3.301), and total motility ( β=-4.932, 95% CI: -7.532--2.331), while positively correlated with sperm concentration ( β=17.288, 95% CI: 9.604-24.973) and DFI ( β=5.226, 95% CI: 3.753-6.699). The results of RCS analysis indicated a significant linear dose-response relationship between age and semen volume ( Pnon-linearity=0.424), showing a decline in semen volume with increasing age ( P<0.001). Non-linear dose-response relationships were observed between age and sperm density ( Pnon-linearity=0.003), sperm progressive motility ( Pnon-linearity<0.001), total motility ( Pnon-linearity<0.001), and DFI ( Pnon-linearity=0.004). Conclusion:In men undergoing assisted reproductive technology for fertility treatment due to non-male factors, age greater than 30 years is significantly associated with a decrease in semen volume, sperm progressive motility, and total motility, as well as an increase in sperm density and DFI.

Objective:To analyze the reflux parameters of patients with gastroesophageal reflux disease (GERD) in upright position, supine position and at 2 h after meals, and to explore the cut-off value, sensitivity and specificity of the reflux parameters in different positions and at 2 h after meals in GERD diagnosis.Methods:From January 2016 to July 2020, 200 GERD patients (GERD group) and 61 non-GERD patients (control group) who visited Huazhong University of Science and Technology Union Shenzhen Hospital (Former Nanshan District People′s Hospital), were selected. All the patients of the two groups received gastroesophageal reflux disease questionnaire (GERDQ), upper gastrointestinal endoscopy, esophageal high resolution manometry and 24 h esophageal pH combined impedance monitoring. T test, non-parametric test and chi-square test were used to compare the related parameters in upright position, supine position and at 2 h after meals between two groups and within each group. Receiver oparative characteristic (ROC) curves of reflux parameters in upright position, supine position and 2 h after meals were drawn to determine the cut-off value, sensitivity and specificity in GERD diagnosis. Results:The proportion of patients with acid reflux in supine position of the control group was higher than that of the GERD group (41.0%, 25/61 vs. 8.50%, 17/200), and the difference was statistically significant ( χ2=36.53, P<0.01). In the control group, the acid reflux time in upright position, number of acid reflux, acid exposure time (AET), longest reflux time and number of weak acid reflux were more than those of in supine position in the same group (6.00 min(2.00 min, 13.50 min) vs. 0.00 min(0.00 min, 1.50 min), 16.00(8.00, 27.00) vs. 1.00(0.00, 3.00), 0.90%(0.33%, 1.88%) vs. 0.00%(0.00%, 0.30%), 2.00 min(1.00 min, 4.00 min) vs. 0.00 min(0.00 min, 1.00 min), 7.00(3.00, 11.00) vs. 1.00(0.00, 2.00), respectively) and the differences were statistically significant ( Z=5.43, 6.61, 5.06, 3.58 and 6.24, all P<0.01). In the GERD group, the acid reflux time, number of acid reflux, AET, longest reflux time and number of weak acid reflux in upright position were higher than those in supine position (51.00 min, (31.00 min, 86.75 min) vs. 8.00 min(1.00 min, 42.00 min), 60.00(48.00, 83.75) vs.6.00(2.00, 19.50), 7.30%(3.90%, 12.10%) vs. 1.50%(0.20%, 6.50%), 7.00 min(4.00, 12.00 min) vs. 4.00 min(1.00 min, 17.00 min), 1.00(0.00, 3.00) vs. 0.00(0.00, 2.00), 7.00(3.00, 12.00) vs. 0.00(0.00, 1.00), respectively) and the differences were statistically significant ( Z=7.92, 11.22, 6.90, 2.56, 5.11 and 11.76, all P<0.05). The acid reflux time, number of acid reflux, AET, longest reflux time and number of weak acid reflux at 2 h postprandial were 3.00 min(2.00 min, 9.00 min), 10.00(5.00, 18.00), 0.90%(0.40%, 1.98%), 1.00 min(0.00 min, 3.00 min), 4.00(1.50, 8.50)and 28.50 min(15.00 min, 54.75 min), 35.00(24.00, 52.00), 8.30%(4.32%, 15.83%), 6.00 min(3.00 min, 11.00 min), 4.00(2.00, 7.25), in the control and GERD groups, respectively, which were significantly higher than those in supine position in the same group ( Z=4.30, 6.33, 5.50, 3.40, 5.71 and 3.76, 9.21, 5.76, 1.97, 10.46, all P<0.05). Among 200 GERD patients, 125 patients had symptoms recorded during the 24 h esophageal pH combined impedance monitoring, the incidence of reflux symptoms in upright position was higher than that in supine position (89.6%, 112/125 vs. 65.6%, 82/125), and the difference was statistically significant ( χ2=20.71, P<0.01). The results of ROC curve analysis showed that the accuracy of acid reflux time in upright position in GERD prediction was the highest, with AUC value of 0.94 and cut-off value of 24.5 min, and the sensitivity and specificity in GERD diagnosis were 81.50% and 95.08%, respectively. The prediction accuracy of acid reflux times in upright position and AET in upright position for GERD was secondary, AUC value both were 0.93 and the cut-off value of the acid reflux number in upright position was 39.5, and the sensitivity and specificity in GERD diagnosis were 84.00% and 95.08%, respectively. The cut-off value of AET in upright position was 2.75%, the sensitivity and specificity in GERD diagnosis were 85.00% and 93.33%, respectively. The AUC value, cut-off value, sensitivity and specificity of AET at 2 h postprandial were 0.91, 4.60%, and 73.49% and 95.00%, respectively. Conclusions:Both GERD patients and non-GERD patients have more reflux in upright position, especially within 2 h after meals. The diagnostic values of acid reflux time in upright position, number of acid reflux, AET and AET 2 h after meals for GERD is high, and the AUC values are all >0.90, which can be used as a more comprehensive basis for the analysis and diagnosis of GERD.

In this research, we compared the visual neuron responses for LGN, A18 and PMLS of old and young cats with extracellular single-neuron recording techniques. We used firing rate vector to characterize information, and response irregularity of cells to evaluate the degeneration of visual characters. Response irregularity is characterized by means of the two coefficients of variation of firing rate vectors: Cv and Cv2. We found that there was no significant change of the response irregularity in LGN areas during the aging process from young to old cats. But in the other two areas, neurons of old cats exhibited significantly larger response irregularity than those of young cats. The result indicated that the information processing function of advanced visual cortex was impaired by aging. This result also provids a reference for the research of the other neuronal system changes during aging process.
Action Potentials ; physiology ; Aging ; physiology ; Animals ; Cats ; Neural Conduction ; Neurons ; physiology ; Visual Cortex ; physiology

Objective To analyze the endoscopic features of patients with the lower esophageal sphincter (LES) and crural diaphragm (CD) separation and explore its significance in hiatus hernia (HH) diagnosis.Methods From January 2011 to June 2012,the data of patients underwent esophagus high resolution manometry (HRM) examination were retrospectively analyzed and of which the patients with LES-CD separation were selected.Patients who received digestive tracts operation or lack of complete endoscopic date were excluded.A total of 93 cases were enrolled for analysis.According to Chicago Criteria,the cases were divided into three groups.There were 21 cases in type Ⅰ group (LES-CD separation＜1 cm),37 cases in type Ⅱ group (LES-CD separation≥l cm and ＜2 cm),35 cases in type Ⅲ group (LES-CD separation≥2 cm).The differences among groups in endoscopic features were analyzed.HRM were taken as golden standard,the sensitivity and specificity of endoscopy in HH diagnosis were analyzed.Measurement data were expressed as x ± s,one-way ANOVA was performed for multi-groups comparisons,LSD method or Dunnett T3 test for pairwise comparisons and chi-square test or Fisher's exact test for two groups' comparisons.Results There were no significant differences in the distance between esophagus gastric junction (EGJ) and foreteeth of the three groups (P＞0.05).Distances between EGJ and diaphragmatic HH pressure impression in type Ⅲ group,type Ⅱ group and type Ⅰ group were (3.57±0.78) cm,(1.89±0.81) cm and (1.14±0.67) cm,respectively and the differences were significant (LSDt=9.26,11.44,3.57; all P＜0.05).The percentage of continuously-open cardia and showing pressure impression of the gastric pouch of type Ⅲ group was higher than that of type Ⅱ group and the percentage of continuously-open cardia and showing pressure impression of gastric poach of type Ⅱ group was higher than that of type Ⅰ group (80.0％,40.5％ and 4.8％,x2 =11.64,29.76 and 8.59; 91.4％,27.0％ and 4.8％,x2 =30.69,40.73 and 4.32' all P＜0.05).The occurrence rates of the His angle blunting and sac of type Ⅲ group were higher than those of typeⅡ group and type Ⅰ group (74.3％ and 77.1％,24.3％ and 24.3％,4.8％ and 4.8％,x2 =17.97 and 25.41,both P＜0.05),there was no significant difference between type Ⅱ group and type Ⅰ group (P＞0.05).The rates of esophagitis of type Ⅲ group and type Ⅱ group were higher than that of type Ⅰ group (71.4％,59.5％ and 14.3％,x2 =17.14 and 11.15,both P＜0.05),there was no significant difference between type Ⅲ group and type Ⅱ group (P＞0.05).There was one case of Barrett's esophagus in each group.The sensitivity and specificity of endoscopy in HH diagnosis were 91.4％ and 81.0％,respectively.Conclusions The distance between EGJ and hiatal pressure impression,continuously-open cardia,the His angle blunting,hiatal pressure impression of the gastric pouch and hernia sac were important indicators of HH endoscopic features.The sensitivity of endosocopy is higher than the spcificity of endosocopy in HH diagnosis.

Objective: To construct and characterize GFP-mfgl 2 fusion protein expression plasmid (pEGFP-mfgl 2) and provide a direct and simplified methodology for primary assessment of the effect of mfgl 2 siRNA on the mfgl 2 gene expression. Methods: mfgl 2 cDNA was amplified from the mfgl 2 cDNA library pBluescript-m166 (pm166) of mouse genomic P1 plasmid and recloned into pEGFP-N2 upstream of GFP gene. The pEGFP-mfgl 2 was analyzed by restriction endonucleases BamH I and Hind III to ensure the orientation and the sequence. This fusion plasmid was then transfected into CHO cells and the fusion protein expression was observed by fluorescent microscope. Rusults: A 1.3 kb long cDNA was obtained. Restriction endonucleases and sequencing assays showed the correct orientation and sequence. After 24-48 hours transfection in CHO cells, the expression of pEGFP-mfgl 2 can be visualized through fluorescent microscope. Conclusion: pEGFP mfgl 2 has been constructed successfully. The recombinant vector can express GFP-mfgl 2 fusion protein. It provides a direct and simplified methodology for primary assessment of the effect of mfgl 2 siRNA on the mfgl 2 gene expression.