Objective The entorhino-hippocampal pathway is the major excitatory input from neurons of the entorhinal cortex on both ipsilateral and contralateral hippocampus/dentate gyrus. This fiber tract consists of the alvear path, the perforant path and a crossed commissural projection. In this study, the histogenesis and development of the various subsets of the entorhino-hippocampal projection have been investigated. Methods DiI, DiO and fast blue tracing as well as anti-calretinin immunocytochemistry were carried out with prenatal and postnatal rats at different ages. Results The alvear path and the commissural pathway started to develop as early as embryonic day (E) 16, while the first perforant afferents reached the stratum lacunosum-moleculare of the hippocampus at E17 and the outer molecular layer of dentate gyrus at postnatal day (P) 2, respectively. Retrograde tracing with DiI identified entorhinal neurons in layer II to IV as the origin of entorhino-hippocampal pathway. Furthermore, anti-calretinin immunocytochemistry revealed transitory Cajal-Retzius (CR) cells in the stratum lacunosum-moleculare of the hippocampus from as early as E16. DiI labeling of entorhinal cortex fibers and combined calretinin-immunocytochemistry showed a close association between CR cells and entorhinal afferents. Conclusion The subsets of entorhino-hippocampal pathway appear in the developmental hippocampus during E16-P2. The temporal and spatial relationship between CR cell and perforant afferent suggests the role of this cell type as a guiding cue for entorhinal afferents at early cortical development.

Acute Disease ; Adolescent ; Adult ; Female ; Hexanes ; poisoning ; Humans ; Male ; Prognosis ; Young Adult

This study was intended to evaluate the effects of hypoxic exposure on gene expression and coordination of cytochrome oxidase (COX) subunits I (COX I) and IV (COX IV) encoded by mtDNA and nDNA respectively in rat cerebral cortex. Male Wistar rats were exposed to hypoxia in a hypobaric chamber simulating high altitude at 5000 m for 2, 5, 15 and 30 d. Control rats were fed outside the hypobaric chamber (the height was 300 m above sea level). Rats were sacrificed and mitochondria from cerebral cortex were isolated by differential centrifugation at each time point. COX I and COX IV proteins in isolated rat cerebral cortex mitochondria were detected by Western blot analysis and mRNA in the cerebral cortex by RT-PCR. The ratios of protein and mRNA were used to estimate the coordinative expression of two subunits. The results showed that COX I mRNA increased significantly at 2 and 5 d, and decreased to the control level at 15 and 30 d; COX IV mRNA remarkably increased at 2, 5 and 15 d, and dropped below the control level at 30 d. The mRNA ratio of COX IV to COX I reached a peak at 15 d, but showed no differences between other time points. The Western blot analysis of COX I and COX IV in isolated rat cerebral cortex mitochondria showed no obvious changes during hypoxic exposure. Our findings demonstrate that hypoxia can affect mRNA expression of COX I and COX IV and their coordination, while protein expression of both subunits are stable and coordinative. This study suggests that the expression of COX I and COX IV proteins during hypoxic exposure is coordinately regulated by post-transcriptional mechanisms.
Animals ; Cerebral Cortex ; metabolism ; Electron Transport Complex IV ; metabolism ; Gene Expression Regulation, Enzymologic ; Hypoxia ; metabolism ; Male ; Mitochondria ; metabolism ; Rats ; Rats, Wistar

Objective Aims to delineate the distribution profile of three isoforms of vesicular glutamate transporter (VGluT), viz. VGluT1-3, and their cellular localization within vestibular nuclear complex (VNC). Methods Brain sections from normal Sprague-Dawley rats were processed immunohistochemically for VGluT detection, employing avidin-biotinylated peroxidase complex method with 3-3'-diaminobenzidine (DAB) as chromogen. Results The whole VNC expressed all of the three transporters that were observed to be localized to the fiber endings. Compared with VGluT1 and VGluT3, VGluT2 demonstrated a relatively homogeneous distribution, with much higher density in VNC. VGluT3 displayed the highest density in lateral vestibular nucleus and group X, contrasting with the sparse immunostained puncta within vestibular medial and inferior nuclei. Conclusion Glutamtatergic pathways participate in the processing of vestibular signals within VNC mainly through the re-uptake of glutamate into synaptic vesicles by VGluT1 and 2, whereas VGluT3 may play a similar role mainly in areas other than medial and inferior nuclei of VNC.

OBJECTIVETo detect the expression of signal transducer and activator of transcription 3 (STAT3) and P-STAT3 in the brain of the APPswe/PS δE9 double transgenic mouse model of Alzhaimer's disease (AD) and investigate their possible role in AD.

METHODSAPPswe/PS δE9 double transgenic mice and control mice were examined for cerebral STAT3 and P-STAT3 expressions using immunothistochemistry.

RESULTSSTAT3 and P-STAT3 were expressed in the different regions of mouse brain. In the transgenic mice and the control mice, the positivity rates of STAT3 were 93.75% and 87.50% in the cerebral cortex, 87.50% and 43.75% in the basal forebrain, 81.25% and 37.50% in the hippocampus, and 62.50% and 0.00% in the cerebellum, respectively, showing significant differences between the mice in the STAT3 expressions in the basal forebrain, hippocampus and cerebellum (P<0.05). The positivity rates of P-STAT3 in the two groups were 0.00% and 0.00% in the cerebral cortex, 68.75% and 0.00% in the basal forebrain, 62.50% and 12.50% in the hippocampus, and 43.75% and 0.00% in the cerebellum, respectively, showing also significant differences in the basal forebrain, hippocampus and cerebellum (P<0.05). The expression of STAT3 was positively correlated with that of P-STAT3 in transgenic AD mice (P<0.05).

CONCLUSIONSTAT3 and P-STAT3 are highly expressed in the basal forebrain, hippocampus and cerebellum in transgenic AD mice and may participate in the pathological process of AD.

Alzheimer Disease ; metabolism ; Animals ; Cerebellum ; metabolism ; Cerebral Cortex ; metabolism ; Disease Models, Animal ; Hippocampus ; metabolism ; Mice ; Mice, Transgenic ; STAT3 Transcription Factor ; metabolism

DNA mismatch repair gene mutS (2.56 kb) was PCR modified and cloned into a secretive prokaryotic expression vector pET32a (+) which carries a N-terminal His.tag + and thioredoxin sequence. MutS protein was expressed with high level after IPTG induction using the strain E. coli AD494(DE3). SDS-PAGE revealed that the expected protein with a molecular weight of 108 kD which is about 35% of the total bacterial proteins is almost soluble. The expected protein was purified directly by immobilized metal (Ni2+) chelation affinity chromatography and the purity is over 90%. MutS protein activity verified using mismatch DNA showed that the expression product can recognize and bind to base-pair mismatch specifically.
Adenosine Triphosphatases ; biosynthesis ; genetics ; isolation & purification ; Bacterial Proteins ; Base Pair Mismatch ; Chromatography, Affinity ; DNA ; metabolism ; DNA Repair ; DNA-Binding Proteins ; Escherichia coli Proteins ; biosynthesis ; genetics ; isolation & purification ; Magnesium ; pharmacology ; Molecular Weight ; MutS DNA Mismatch-Binding Protein ; Recombinant Proteins ; biosynthesis

OBJECTIVETo investigate the presence of β-amyloid peptide (Aβ) deposition in the cerebellum and the expression of related miRNAs in the cerebellum of a mouse model of Alzheimer disease.

METHODSTwelve 12-month-old APPswe/PSδE9 double transgenic mice and 12 wild-type C57 mice were sacrificed and the brain tissues were taken for examination. The right hemisphere was stained with Congo red to observe the deposition of amyloid substances, and from the left hemisphere, the hippocampus and the cerebellum were dissected for detecting the expression of miRNA-135a-5p, miRNA-298-5p, miRNA-466b-3p and miR-669f-3p using real-time PCR.

RESULTSCongo red staining revealed the presence of Aβ deposition in both the hippocampus and the cerebellum of the transgenic mice but not in the control mice. Real-time PCR showed a significantly lower expression of the 4 miRNAs in the hippocampus in the transgenic mice than in the control mice (P<0.05). The expression of miRNA-135a-5p, miRNA-298-5p, and miR-669f-3p in the cerebellum was significantly lower in the transgenic mice than in the control mice (P<0.05). The expression of miRNA-298-5p and miR-669f-3p in the hippocampus was significantly lower than that in the cerebellum of the transgenic mice (P<0.05).

CONCLUSIONβ deposition also occurs in the cerebellum of APPswe/PSδE9 double transgenic mice, and its formation might be related to the down-regulation of miRNA-135a-5p, miRNA-298-5p, and miR-669f-3p.

Alzheimer Disease ; metabolism ; Amyloid beta-Peptides ; metabolism ; Animals ; Cerebellum ; metabolism ; Disease Models, Animal ; Hippocampus ; metabolism ; Mice ; Mice, Inbred C57BL ; Mice, Transgenic ; MicroRNAs ; metabolism

Objective:To investigate the effect of early gradual diet on reducing delirium in elderly patients with hip arthroplasty.Methods:From January 2018 to January 2020, 74 cases of hip arthroplasty patients aged over 65 years old who were treated in the Third Affiliated Hospital of Sun Yat-sen University were selected as the observation objects. They were randomly divided into experimental group and control group with 37 cases in each group. The experimental group was given early gradual diet on the basis of routine postoperative care, while the control group was given routine postoperative diet on the basis of routine postoperative care. The incidence of postoperative delirium, Pittsburgh Sleep Quality Index (PSQI), patient satisfaction rate, average hospitalization days and average hospitalization expenses were used to evaluate the effect of early gradual diet on reducing delirium in elderly patients with hip arthroplasty.Results:The incidence of delirium in the experimental group was 2.70% (1/37) and 16.22% (6/37) in the control group, the difference was statistically significant ( χ2 value was 3.945, P<0.05); the hospitalization days of the experimental group were (10.68±5.13) d, (13.62±7.19) d in the control group. The difference of hospitalization days was statistically significant ( t value was 2.877, P<0.01). The incidence of difficulty in falling asleep and the satisfaction rate of the experimental group were 8.11% (3/37) and 94.59% (35/37) respectively, and those in the control group were 29.73% (11/37) and 78.38% (29/37) respectively, and the differences were statistically significant ( χ2 value was 5.638, 4.163, P<0.05). Conclusions:Early gradual diet after operation can reduce the incidence of delirium in elderly patients with hip arthroplasty, shorten the average hospitalization days, reduce the incidence of difficulty in falling asleep, improve patients' satisfaction, and help patients to pass through the perioperative period more safely and comfortably.

OBJECTIVETo investigate the role of interleukin-17 (IL-17) in alveolar fluid clearance in mice with acute lung injury (ALI) and explore the possible mechanism.

METHODSSixteen IL-17-knockout mice and 16 wild-type mice were both randomized for intratracheal instillation of PBS (control) on lipopolysaccharide (LPS) to induce ALI. Forty-eight hours after the treatments, the wet-dry ratio (W/D) of the lungs, IL-8 in the bronchoalveolar lavage fluid (BALF) and histopathological changes of the lung tissues were examined. The expressions of epithelial sodium channel α subunit (α-ENaC) was detected with Western blotting and liver kinase B1 (LKB1) was detected with immunohistochemistry.

RESULTSCompared with wild-type mice treated with LPS, IL-17 knockout mice showed significantly decreased W/D of the lungs (9.739∓3.3 vs 5.351∓0.56) and IL-8 level in the BALF (67.50∓7.33 vs 41.00∓3.16 pg/mL) following LPS challenge. Pathological examination revealed reduced alveolar edema fluid aggregations and lower lung injury score in IL-17 knockout mice with also higher expression levels of ENaC and LKB1 compared with the wild-type mice.

CONCLUSIONKnocking out IL-17 in mice not only alleviates inflammation of the lung tissue following ALI but also reduces the loss of ENaC protein and promotes alveolar fluid clearance, mechanism of which is probably associated with LKB1.

Acute Lung Injury ; metabolism ; Animals ; Bronchoalveolar Lavage Fluid ; chemistry ; Epithelial Sodium Channels ; metabolism ; Gene Knockout Techniques ; Interleukin-17 ; genetics ; metabolism ; Interleukin-8 ; metabolism ; Lipopolysaccharides ; Lung ; pathology ; Mice ; Protein-Serine-Threonine Kinases ; metabolism

OBJECTIVETo assess the larvicidal effects of recombinant Escherichia coli expressing scorpion neurotoxin AaIT or Bacillus thuringiensis subsp israelensis (B.t.i) toxin Cyt2Ba against the second instar larvae of Culex pipiensquinquefasciatus and Aedes albopictus and compare different formulations for their larvicidal effects.

METHODSThe AaIT- or Cyt2Ba-coding sequences were cloned into pET28a(+) and the recombinant plasmids were transformed into E. coli BL21(DE3). After induction with IPTG, the recombinant proteins expressed by the recombinant E. coli were detected and identified by SDS-PAGE and Western blotting, respectively. The larvicidal activity of the bacterial suspension was tested at different concentrations against mosquitoes. The effective engineered bacteria were prepared into dry powder with different formulations, and their larvicidal activity was tested.

RESULTSAaIT and Cyt2Ba proteins were successfully expressed in E. coli. The recombinant AaIT protein showed no virulence to the mosquito larvae. The suspension of the recombinant E. coli expressing Cyt2Ba protein exhibited a stronger killing effect on Aedes albopictus larvae than on Culex pipiens quinquefasciatus larvae at 48 h (P<0.001) with LCof 3.00×10cells/mL and 1.25×10cells/mL, respectively. The dry powder of the engineered bacteria formulated with yeast extract, wheat flour or white pepper powder at the mass ratio of 1:1 showed the strongest killing effect on mosquito larvae (P=0.044), and the formulation with white pepper powder produced a stronger killing effect than formulations with yeast extract or wheat flour (P=0.002).

CONCLUSIONThe B.t.i Cyt2Ba protein expressed in E. coli BL21(DE3) shows a good larvicidal activity against mosquitoes, and appropriate formulations of the engineered bacteria can enhance its efficiency in mosquito control.