Objective To study the condition and method of hydrodynamics-based transgene(HDT) in rat fatty liver. Methods Inject different dosages and concentrations of green fluorescent protein plasmid pEGFP-C1 at different speeds, then collect 4 rats' liver leaves at different time points after injection and prepare their frozen section, finally observe and quantify the GFP expression with fluo rescence microscope at 488 nm excitation wavelength. Results Plasmid pEGFP-C1 concentration 33mg/L, injection speed 2ml/s, injection volume 8.5%of rat body weight, injected plasmid. After 6 hours of injection, GFP-positive cells rate of pedicel leaf is about 18%, left leaf about 14%, middle leaf about 12.5%;R3ight leaf about 10% and tail leaf about 8%. GFP begin to gradually reduce since 24 hours, until 72 hours almost no GFP-positive cells were checked in all liver leaves. Conclusion Hydrodynamics-based transgene can be applied to rat fatty liver, the appropriate conditions of this method are 33mg/L plasmid concentration, 8.5% rat avoirdupois, 2ml/s injection speed, and the suitable time to observe the proportion of GFP-positive cells is 6-24 hours after gene injection.

At present,intranasal delivery has entered clinical trial stage.In recent years,how to imroving the efficiency of intranasal delivery has been extensively investigated.This article briefly reviews some factors that impact the targeting of intranasal delivery and the drug concentration in the central nervous system.

OBJECTIVE:To reduce the error rate of inventory in the automated pharmacy of our hospital. METHODS:Activi-ties were designed and implemented by the management method of quality control circles(QCC)-PDCA(Plan,Do,Check and Ac-tion)cycle. The reasons for the errors of inventory in the automated pharmacy were analyzed to investigate and implement counter-measures. Visible and invisible achievements were evaluated,and then standardized processes were made. RESULTS:The errors of inventory in the automated pharmacy mainly included those of drug dispensing,drug shelving,drug return sheet,automatic medi-cine dispensing machine and system. In view of the above reasons,relevant standards were formulated and performed,including the process of warehouse-out check and shelving of drugs,drug dispensing process for the automated pharmacy,the process of sec-ondary check,etc. After the implementation of the activities,the error rate of inventory in the automated pharmacy reduced from 9.17% to 3.77%,which was visible achievement;and the above-mentioned standardized processes could ensure continuous run-ning of PDCA cycle. The practice,sense of responsibility,communication and coordination of management members in PDCA cy-cle namely invisible achievements were improved to some extent. CONCLUSIONS:The management method of QCC-PDCA cycle is feasible in reduction of the error rate of inventory in the automated pharmacy,which can provide a reference for automated phar-macy management.

Objective To study the effect of intranasal nerve growth factor (NGF) on the expression of amyloid-β,peptide (Aβ) in the central nervous system in rats with traumatic brain injury (TBI).Methods Eighty rats were randomly divided into sham(n =26),control(n =27) and treatment group (n =27 ).They were subjected to the modified Feeney' s weight-drop model.The treatment group was treated with NGF administered by nasal route,and the control group was given phosphate-buffered saline (PBS).Beam walking and Morris water maze test were performed in the three groups.The concentration of Aβ40 and Aβ42 in the injured ipsilateral hippocampus was elevated by ELISA measurement.Immunohistochemistry was used to detect the amyloid precursor protein (APP) positive cells near the region of injury in the hippocampus in rats after TBI.Results NGF group traversed the beam significantly quicker (s) than control group ( 19.00 + 6.99 vs 27.33 ± 7.39 respectively,F2,15 =12.87,P =0.028 ).Morris water maze performance revealed that mean time of latency in the NGF group was significant shorter than vehicle group,and significant memory retention in NGF group as evidenced by a greater percentage of the 60 s allotted time spent in the target quadrant (45.82％ ± 11.15％ vs 33.99％ ± 3.46％,F2,15 =6.814,P=0.037),as well as the number crossing of the former site of the removed platform in NGF group was significant more than control group (8.60 ±2.73 vs 3.60 ±2.06,F2,15 =5.346,P =0.04).The Aβ42 level in control group was increased significantly higher than NGF group as indicated by ELISA measurements.While the Aβ40 level did not have similar shown.Immunohistochemical staining showed that APP level had significant differences among three groups ( F2,15 =8.672,P =0.003).The APP level in NGF group did not alter with control group.Conclusion Intranasal administration of NGF can regulate Aβ42 overproduction,improve the motor and cognitive function after brain injury in rats.

Objective To investigate the protective role of human serum albumin in treatment of monocyte-inducible C-type lectin (Mincle)-associated neuroinflammation in subarachnoid hemorrhage (SAH) rats and its underlying mechanisms.Methods Vascular perforation model was used to induce SAH.Ninety-two male SD rats were randomly assigned to sham (n =23),vehicle (n =23),low-dose albumin (0.63 g/kg,n =23) and high-dose albumin (1.25 g/kg,n =23) groups.Saline and albumin were intravenously injected into rats two hours after surgery.Modified Garcia scale was employed to assess neurological functions.Iba-1 and myeloperoxidase (MPO) staining was used to examine the activation of microglial cells and infiltration of neutrophils.Real-time PCR was applied to determine the changes of IL-1β,inducible nitric oxide synthase,CD11b,monocyte chemoattractant protein-1,cytokine-induced neutrophil chemoattractant-1 and CXC motif chemokine ligand-2 mRNA levels.Co-immunoprecipitation was conducted to assess the binding ability of albumin with Mincle.Immunoblotting was carried out to evaluate protein levels of Minlce,Syk and p-Syk.SAH severity measurement was performed before conductions of all the experiments.Results SAH severity scores were 11.4 ± 1.6,12.8 ±2.5 and 11.2 ±3.2 in the vehicle,low-dose albumin and high-dose albumin groups,respectively,without statistically significant difference among groups (F =0.694,P =0.516).Neurological score was 7.5 ± 2.9 in the vehicle group,while the low-dose albumin (14.6 ± 2.2) and high-dose albumin groups (13.6 ± 2.7) exhibited better neurological perfomance (P ＜ 0.01).Immunostaining showed that albumin significantly inhibited the activation of microglia,and reduced the percentage of MPO positive cells from 20.7％ ± 1.9％ in the vehicle group to 12.1％ ±2.1％ in the low-dose albumin group and 9.8％ ±0.9％ in the high-dose albunin group (F =32.216,P =0.001).mRNA levels of pro-inflammatory cytokines and chemotactic factors were also suppressed by albumin (P ＜ 0.05).The results of co-immunoprecipitation displayed that albumin could directly bind Mincle and disrupt the association between Mincle and SAP130.Immunoblotting demonstrated that albumin depressed the protein levels of Mincle,Syk and p-Syk.Conclusion Human serum albumin can inhibit Mincle/Syk-induced neuroinflammation via directly binding Mincle receptor in SAH rats.

BACKGROUND:Estrogen signaling pathway for interaction between aromatase and estrogen-related receptor may exist in bone marrow mesenchymal stem cel s, which is used for regulating biological activity of bone marrow mesenchymal stem cel s. OBJECTIVE:To observe the expression of aromatase and estrogen-related receptors in adult bone marrow mesenchymal stem cel s during osteogenic differentiation. METHODS:Bone marrow mesenchymal stem cel s were respectively cultured in low-glucose DMEM medium (control group) and osteogenic induction medium (induction group). Cel proliferation and calcium deposition were determined by MTT assay and alizarin red staining, respectively. The expression of aromatase, estrogen receptorα, estrogen receptorβ, and estrogen-related receptorαduring osteogenic differentiation were determined by real-time PCR and western blot analysis. Estradiol levels in supernatants and lysates were detected by ELISA method. RESULTS AND CONCLUSION:In the induction group, the proliferation ability of bone marrow mesenchymal stem cel s was the strongest at 72 hours of culture;while there were a great amount of calcium nodules formed at 21 days of culture. Results from PCR and western blot assay showed that the expression of aromatase and estrogen receptorαwas improved in the induction group, but the expression of estrogen-related receptorαwas inhibited. There was no difference in the expression of estrogen receptorβbetween the two groups. ELISA results indicated that the level of estradiol in the supernatant of induction group was the highest. These findings indicate that aromatase, estrogen receptorα, estrogen receptorβand estrogen-related receptorαare al involved in osteogenesis of bone marrow mesenchymal stem cel s. Moreover, estradiol can be synthesized and secreted in bone marrow mesenchymal stem cel s, and most likely, promote the osteogenic differentiation of bone marrow mesenchymal stem cel s by related receptor pathway.

White matter lesion is a major subtype of cerebral small vessel disease. Its pathophysiology and mechanism remain unclear. Because the risk factors often coexist in clinical research, it is difficult to judge the relationship between certain risk factors and white matter injury. Moreover, due to the differences in animal and human brain tissue structure, there is currently a lack of reproducible animal models of white matter lesions. Therefore, establishing a practical animal model and further exploring the pathogenesis and risk factors of white matter lesions from the basic research level is crucial for the preclinical study of the treatment of white matter damage. This article reviews the characteristics, optimization measures, and application prospects of the white matter lesion models.

BACKGROUND:The sclerotic zone in the femoral head is an important imaging feature in the progression of steroid-induced femoral head necrosis,which is associated with disease prognosis.Peroxisome proliferator-activated receptor γ coactivator 1α(PGC-1α)has been shown to possess biological activities such as osteogenesis,angiogenesis and anti-mitochondrial apoptosis,which may be closely related to bone repair of steroid-induced femoral head necrosis. OBJECTIVE:To screen for the differential proteins in the sclerotic zone of steroid-induced osteonecrosis of the femoral head versus the normal zone,to screen for hub proteins in the sclerotic zone,and to verify the differential expression of hub proteins in the femoral head specimens following steroid-induced femoral head necrosis,and to to explore the repair pattern of the sclerotic zone following steroid-induced femoral head necrosis. METHODS:Femoral head samples were collected from patients with steroid-induced osteonecrosis of the femoral head receiving total hip arthroplasty.The differentially expressed genes in the sclerotic zone and the normal zone were screened by Tandem Mass Tags and analyzed by GO and KEGG signaling pathways to construct a protein-protein interaction network and screen hub genes.In addition,the expression of hub genes in the sclerotic zone was verified by immunohistochemistry and western blot. RESULTS AND CONCLUSION:Quantitative protein profiling by Tandem Mass Tags revealed that 609 proteins were significantly differentially expressed(Log2FC＞1.20,Log2FC＜0.84 and P＜0.05)in the sclerotic zone of the femoral head compared with the normal zone,of which 290 proteins were upregulated and 319 proteins were downregulated.The GO and KEGG pathway enrichment analyses revealed that among the top 10 enriched pathways,Wnt signaling pathway and life-cycle regulatory pathway were closely related to bone repair;in the life-cycle regulatory pathway,PGC-1α was one of the important proteins.In addition,western blot results verified the low expression of PGC-1α and NRF1 in the sclerotic zone and high expression of Cleaved Caspase-3 in the sclerotic zone compared with the normal zone of steroid-induced femoral head necrosis specimens.Light microscopic immunohistochemical results showed the distribution of PGC-1α,NRF1 and Cleaved Caspase-3 positive expression in the sclerotic and normal zones in the femoral head tissue specimens,indicating the presence of their expression in bone trabeculae,osteoblasts and bone marrow.In contrast,the brown area of the sclerotic zone of femoral head necrosis stained darker and showed more obvious expression of Cleaved Caspase-3.To conclude,in the sclerotic zone of steroid-induced femoral head necrosis,biological behaviors including activation of osteogenesis-related pathways such as Wnt and oxidative apoptosis characterized by low expression of PGC-1 are observed.Low expression of PGC-1α in the sclerotic zone of steroid-induced femoral head necrosis may be associated with the activation of oxidative apoptosis.

The many-banded krait, Bungarus multicinctus, has been recorded as the animal resource of JinQianBaiHuaShe in the Chinese Pharmacopoeia. Characterization of its venoms classified chief phyla of modern animal neurotoxins. However, the evolutionary origin and diversification of its neurotoxins as well as biosynthesis of its active compounds remain largely unknown due to the lack of its high-quality genome. Here, we present the 1.58 Gbp genome of B. multicinctus assembled into 18 chromosomes with contig/scaffold N50 of 7.53 Mbp/149.8 Mbp. Major bungarotoxin-coding genes were clustered within genome by family and found to be associated with ancient local duplications. The truncation of glycosylphosphatidylinositol anchor in the 3'-terminal of a LY6E paralog released modern three-finger toxins (3FTxs) from membrane tethering before the Colubroidea divergence. Subsequent expansion and mutations diversified and recruited these 3FTxs. After the cobra/krait divergence, the modern unit-B of β-bungarotoxin emerged with an extra cysteine residue. A subsequent point substitution in unit-A enabled the β-bungarotoxin covalent linkage. The B. multicinctus gene expression, chromatin topological organization, and histone modification characteristics were featured by transcriptome, proteome, chromatin conformation capture sequencing, and ChIP-seq. The results highlighted that venom production was under a sophisticated regulation. Our findings provide new insights into snake neurotoxin research, meanwhile will facilitate antivenom development, toxin-driven drug discovery and the quality control of JinQianBaiHuaShe.