BACKGROUND:Meningeal lymphatic vessels can drain cerebral spinal fluid and amyloid β-protein,promoting T lymphocyte to transport and home to deep cervical lymph nodes.A simple,quick and definite method of dural separation and accurate localization of deep cervical lymph nodes can provide strong support for the study of neurodegenerative diseases.OBJECTIVE:To establish a convenient and practical method for exfoliating dural and deep cervical lymph nodes.METHODS:ICR mice,3 months old,were taken,anesthetized and injected with Evans blue and tracer in the occipital pool for localizing deep cervical lymph nodes.A midline incision of about 3 cm in length was made about 5 mm above the clavicle,the superficial fat and fascia were bluntly separated,and the lateral sternocleidomastoid muscle was pulled to expose the deep cervical lymph nodes,which were removed under a stereomicroscope and frozen at-80℃.Subsequently,the mouse head was cut and the skin and muscles of the head were separated to expose the entire skull structure.The skull and brain tissue were separated from the foramen magnum along the lower parietal bone with scissors,and the complete skull top was obtained.The skull was sequentially fixed in 40 g/L paraformaldehyde solution for 24 hours,120 g/L paraformaldehyde for 24 hours,and 120 g/L paraformaldehyde for 10,20,30,and 40 minutes,and the dural structure was stripped.The drainage capacity of meningeal lymphatic vessels and deep cervical lymphatic vessels was verified by tracer,and the meningeal lymphatic vessels were identified by the lymphatic vessel endothelial hyaluronan receptor 1 using the immunofluorescence method.RESULTS AND CONCLUSION:(1)Obvious blue staining was observed in deep cervical lymph nodes 15 minutes after Evans blue staining.(2)The skull was sampled and fixed in 120 g/L paraformaldehyde for 24 hours,resulting in a less tight connection between the dura mater and the skull,and easier stripping of the dural structures with an intact shape.The dura mater fixed at 120 g/L concentration was more resilient and remained more intact during peeling compared with the conventional 40 g/L concentration;120 g/L paraformaldehyde fixed meninges for a short time,and 30-40 minutes was preferred.(3)The frozen section of deep cervical lymph nodes showed the presence of the tracer,complete meningeal lymphatic vessels were visible in the dura mater,and the tracer was observed at the tail of lymphatic vessels.Immunofluorescence staining for endothelial hyaluronan receptor 1 was positive in the deep cervical lymph nodes and dural lymphatics.In summary,the best peeling concentration and time is 120 g/L paraformaldehyde fixed for 24 hours.At this concentration,the dura mater has a stretched morphology,a better toughness,and is more intact after peeling,which is conducive to later use.Verified by Evans blue,tracers and immunofluorescence,deep cervical lymph nodes are located accurately,which can be used as a basis for the study of various neurodegenerative diseases.

BACKGROUND:Meningeal lymphatic vessels can drain cerebral spinal fluid and amyloid β-protein,promoting T lymphocyte to transport and home to deep cervical lymph nodes.A simple,quick and definite method of dural separation and accurate localization of deep cervical lymph nodes can provide strong support for the study of neurodegenerative diseases.OBJECTIVE:To establish a convenient and practical method for exfoliating dural and deep cervical lymph nodes.METHODS:ICR mice,3 months old,were taken,anesthetized and injected with Evans blue and tracer in the occipital pool for localizing deep cervical lymph nodes.A midline incision of about 3 cm in length was made about 5 mm above the clavicle,the superficial fat and fascia were bluntly separated,and the lateral sternocleidomastoid muscle was pulled to expose the deep cervical lymph nodes,which were removed under a stereomicroscope and frozen at-80℃.Subsequently,the mouse head was cut and the skin and muscles of the head were separated to expose the entire skull structure.The skull and brain tissue were separated from the foramen magnum along the lower parietal bone with scissors,and the complete skull top was obtained.The skull was sequentially fixed in 40 g/L paraformaldehyde solution for 24 hours,120 g/L paraformaldehyde for 24 hours,and 120 g/L paraformaldehyde for 10,20,30,and 40 minutes,and the dural structure was stripped.The drainage capacity of meningeal lymphatic vessels and deep cervical lymphatic vessels was verified by tracer,and the meningeal lymphatic vessels were identified by the lymphatic vessel endothelial hyaluronan receptor 1 using the immunofluorescence method.RESULTS AND CONCLUSION:(1)Obvious blue staining was observed in deep cervical lymph nodes 15 minutes after Evans blue staining.(2)The skull was sampled and fixed in 120 g/L paraformaldehyde for 24 hours,resulting in a less tight connection between the dura mater and the skull,and easier stripping of the dural structures with an intact shape.The dura mater fixed at 120 g/L concentration was more resilient and remained more intact during peeling compared with the conventional 40 g/L concentration;120 g/L paraformaldehyde fixed meninges for a short time,and 30-40 minutes was preferred.(3)The frozen section of deep cervical lymph nodes showed the presence of the tracer,complete meningeal lymphatic vessels were visible in the dura mater,and the tracer was observed at the tail of lymphatic vessels.Immunofluorescence staining for endothelial hyaluronan receptor 1 was positive in the deep cervical lymph nodes and dural lymphatics.In summary,the best peeling concentration and time is 120 g/L paraformaldehyde fixed for 24 hours.At this concentration,the dura mater has a stretched morphology,a better toughness,and is more intact after peeling,which is conducive to later use.Verified by Evans blue,tracers and immunofluorescence,deep cervical lymph nodes are located accurately,which can be used as a basis for the study of various neurodegenerative diseases.

Objective:To establish a method for the determination of oleanolic acid in She medicine Clematis florida var. plena. Methods:The HPLC method was carried on a Lanbo-Kromasil C18 chromatography column using acetonitrile-water ( 86∶ 14 ) as the mobile phase. The detection wavelength was 205 nm, the flow rate was 1. 0 ml·min-1 , and the column temperature was 25 ℃. Re-sults:The calibration curve of oleanolic acid was linear within the range of 0. 52-15. 60μg (r=0. 999 9). The average recovery of oleanolic acid was 92. 13%(RSD=3. 0%, n=9). Conclusion:The method is simple, rapid, accurate and reproducible, which can be used as an ideal method for the quality control of She medicine Clematis florida var. plena. .

Objective To investigate the features of the auditory mismatch response (MMR) elicited from the preschoolers and adults.Methods 9 preschoolers aged 3-6 and 8 adults were elicited and measured MMR to speech sounds (/bal/, /pal/) using the Oddball paradigm.Results The response was typical mismatch negativity in adults, and was slow positive waves with larger amplitude in the preschoolers.MNOVA results showed that there were significantly differences between the 2 groups, said the latency of MMRs was significantly longer and the amplitude was larger in the preschoolers than in the adults (P<0.05). Conclusion Stable MMRs with distinct characters in preschoolers and adults have been obtained respectively.

Objective To compare the di-syllabic tone between hearing-impaired children and normal-hearing children. Methods 120 hearing-impaired children and 36 normal-hearing children aged 3~5 years were asked to read aloud 32 word with 16 kinds of di-syllabic tone. Results All the normal-hearing children read all the words correctly, and the hearing-impaired children faulted in pronouncing as their peers. The correct incidence improved in 4-year old hearing-impaired children compared with that of the 3-year old, but it was not significant different between the 4- and the 5- year old. There was not significant difference of the high and level tone between the hearing-impaired children and normal-hearing children. However, the rising, falling- rising and falling tones were significantly different between the two groups. Conclusion The hearing-impaired children aged 3~5 years delay to speech words with di-syllabic tone, especially for those with rising, falling-rising and falling tones.

Objective To evaluate the academic level of Chinese Journal of Internal Medicine by analyzing its citation status by using bibliometrics method.Methods The distribution of articles published in Chinese Journal of Internal Medicine from January,2005 to December,2011 indexed by Chinese Science Citation Database (CSCD) was analyzed.Results A total of 2809 articles were published in Chinese Journal of Internal Medicine between January,2005 and December,2011.Among them,832 (29.62％) articles were cited for totally 1993 times.There were 14 authors whose total citation number in 2005-2011 was ≥ 10 times.Authors of the cited articles came from 25 provinces,autonomous regions and municipalities.The regions from where the articles had the highest citation were Beijing (341 articles),Shanghai (87 articles),Guangdong (64 articles),Jiangsu (45 articles) and Zhejiang (43 articles).The medical institutions with the highest citation were Peking Union Medical College Hospital of Peking Union Medical College and Chinese Academy of Medical Sciences (205 times),Peking University People's Hospital (77 times),and Chinese PLA General Hospital (76 times).Conclusions Quite a few high level academic papers had been published in Chinese Journal of Internal Medicine in recent years.Chinese Journal of Internal Medicine plays a good role in enhancing academic exchange.

Objective To outline the research fronts of ten major sub-specialties in internal medicine.Methods Based on their impact factor scores and the proportion of the journals of 10 subspecialties (endocrinology & metabolism,cardiac & cardiovascular systems,hematology,infectious diseases,nephrology,gastroenterology & hepatology,respiratory system,rheumatology,critical care medicine,clinical neurology) in Journal Citation Report (JCR),and careful consulation of expert clinicians,we identified 50 journals.Their bibliographic records (including references) published in 2011 were downloaded,and the frequency of the references (citations) in each sub-specialty was counted and the highly cited records were extracted.We performed a clustering analysis according to the co-cited times among any pairs of the highly cited records.To tag each cluster of highly cited records,we browsed the titles and abstracts of all highly cited records in the same cluster,and concluded the main topics of each cluster.Finally,we extracted the current published papers devoted to particular cluster by some clustering analysis indicators.The clusters of highly cited records were considered as the intelligence base,and the main topics in current papers which citing these highly cited papers were considered as research fronts.Results Totally 50 journals on 10 sub-specialties in internal medicine were identified.A total of 202 highly cited papers,38 clusters (knowledge bases),and 152 corresponding current citing papers presenting the research fronts were selected.Conclusions We confirm and present research fronts in 10 major sub-specialties of internal medicine.This study provides a synchronic structure of contemporary research activities in internal medicine sub-specialties.

Objective To analyze the articles and citation published in the Chinese Journal of Internal Medicine from 2008 to 2010,in order to investigate the influence factors of impact factor (IF).Methods All articles published in the Chinese Journal of Internal Medicine covered by Chinese Medical Citation Index(CMCI) from 2008 to 2010 were searched and downloaded.Some article related attributions were manual added and their influences to IF were analyzed.Results A total of 1164 academic papers were published in the journal in 3 years,with 9.95 references per paper.The total citation was 1029 times,with 0.93 time per paper and 0.31 time per page.Among them,736 articles were not cited,accounting for 63.2％.Original articles,pure clinical articles had good citation output.For fund supporting,the citation of articles without fund was higher than those with fund.The articles on hematology,cardiology and gastroenterology accounted the most,while ICU,emergency and nephrology were the least internal medicine reported specialties.Although there exists citation difference among different subjects,for considering the absolute values,neurology/psychiatry (0.73 time per paper),cardiology (0.65 time per paper) and gastroenterology (0.54 time per paper) had better citation output,while hematology,basic research and rheumatology had no good performance to IF.Conclusion We should further strengthen acquisition and dissemination of excellent articles,reduce the number of non-cited paper,expand periodical visibility,and provide a quick and convenient way of literature reading.

OBJECTIVEFactor VIII( FVIII) gene knockout mouse model was established for further study on the treatment of hemophilia A.

METHODSExons 16-19 of the mouse FVIII gene were knocked out by ET clone, ES homologous recombination and tetraploid embryo compensation technology. PCR, reverse transcriptase-PCR(RT-PCR) and immunohistochemistry were used to detect the transcription and translation pattern of FVIII. The phenotype of the knockout mice was analyzed by examining the activated partial thromboplastin time (APTT) and FVIII activity (FVIII:C).

RESULTSPCR, RT-PCR and immunohistochemistry confirmed that FVIII was deficient in the FVIII gene knockout mouse. The APTT results showed that FVIII-deficient mouse plasma had a prolonged clotting time compared to normal mouse plasma. The FVIII:C in heterozygous, hemizygous and homozygous mice was 80%, 8% and 10% of that in normal mice, respectively.

CONCLUSIONThe phenotype of the FVIII gene knockout mouse appears grossly similar to that of human with hemophilia A. Establishment of this model may promote the development of new technologies of treatment to hemophilia A.

Animals ; Disease Models, Animal ; Embryo, Mammalian ; Factor VIII ; genetics ; metabolism ; Female ; Hemophilia A ; genetics ; metabolism ; physiopathology ; Humans ; Male ; Mice ; Mice, Inbred ICR ; Mice, Knockout ; Partial Thromboplastin Time