Tumor cell-based glycolytic metabolism and single-cell sequencing of urinary exfoliated cells for the diagnosis and molecular profiling of urothelial carcinoma / 中华病理学杂志

Objective: To explore the diagnostic values of HK2 testing and single-cell sequencing in the urothelial carcinoma (UC). Methods: The qualified urine specimens of 265 suspected UC patients or postoperative patients from the Cancer Hospital of Chinese Academy of Medical Sciences, Beijing, China were collected. Both exfoliative cytology and HK2 testing were performed on clinically suspected UC or postoperative patients. The performance of diagnostic cytology and HK2, including consistency, sensitivity, specificity, positive predictive value and negative predictive value, was evaluated based on histopathological, clinical and imaging diagnosis. Isolated HK2 metabolically abnormal cells were subject to single-cell sequencing to verify the reliability of HK2 detection performance and to explore the molecular characteristics of UC. Results: The concordance rate of HK2 testing and cytology for detecting UC was 90.3% (102/113, Kappa=0.604). Compared with cytology, the sensitivity of HK2 was significantly higher (85.2% versus 75.6%, P=0.024). The detection sensitivity of combined HK2 testing and cytology was increased to 91.1%. HK2 testing was significantly more sensitive than cytology for diagnosing UC in the upper urinary tract (81.8% versus 65.5%, P=0.022). It was also more sensitive than cytology for diagnosing early-stage UC (82.6% versus 69.5%, P=0.375) and low-grade UC (69.6% versus 47.8%, P=0.125). Single-cell sequencing of the ten patients, whose samples were positive for HK2, demonstrated highly concordant copy number variations (CNVs) in tumor cells from the same UC patient, with heterogeneity in CNV profiles among different patients. Deletion of chromosome 8p was found in 3 of the 4 urine samples of renal pelvis UC. The 2 patients with benign lesions had no CNVs in all sequenced cells. Conclusions: The test for abnormal urinary glycolytic HK2 metabolism can assist urine cytology to improve the sensitivity of UC diagnosis, and it provides a novel and reliable approach for early detection of upper urinary tract UC and lower grade UC. Meanwhile, this study has preliminarily revealed the feasibility of single-cell sequencing in urinary samples, which is expected to improve the diagnostic specificity of HK2 testing.

Effects of ring finger and tryptophan-aspartic acid 2 on dendritic spines and synapse formation in cerebral cortex neurons of mice / 南方医科大学学报

OBJECTIVE@#To clarify the functional effects of differential expression of ring finger and tryptophan-aspartic acid 2 (RFWD2) on dendritic development and formation of dendritic spines in cerebral cortex neurons of mice.@*METHODS@#Immunofluorescent staining was used to identify the location and global expression profile of RFWD2 in mouse brain and determine the co-localization of RFWD2 with the synaptic proteins in the cortical neurons. We also examined the effects of RFWD2 over-expression (RFWD2-Myc) and RFWD2 knockdown (RFWD2-shRNA) on dendritic development, dendritic spine formation and synaptic function in cultured cortical neurons.@*RESULTS@#RFWD2 is highly expressed in the cerebral cortex and hippocampus of mice, and its expression level was positively correlated with the development of cerebral cortex neurons and dendrites. RFWD2 expression was detected on the presynaptic membrane and postsynaptic membrane of the neurons, and its expression levels were positively correlated with the length, number of branches and complexity of the dendrites. In cultured cortical neurons, RFWD2 overexpression significantly lowered the expressions of the synaptic proteins synaptophysin (P < 0.01) and postsynapic density protein 95 (P < 0.01), while RFWD2 knockdown significantly increased their expressions (both P < 0.05). Compared with the control and RFWD2-overexpressing cells, the neurons with RFWD2 knockdown showed significantly reduced number of dendritic spines (both P < 0.05).@*CONCLUSION@#RFWD2 can regulate the expression of the synaptic proteins, the development of the dendrites, the formation of the dendritic spines and synaptic function in mouse cerebral cortex neurons through ubiquitination of Pea3 family members and c-Jun, which may serve as potential treatment targets for neurological diseases.

Effect of CDK1 Interferes with the Regulation of PLK1, Aurora B and TRF1 on the Proliferation of Leukemia Cells / 中国实验血液学杂志

OBJECTIVE@#To investigate the clinical characteristics, diagnosis and treatment of 1 case EBV negative extranodal NK/T cell lymphoma (ENKTL) patients.@*METHODS@#The clinical manifestations, diagnosis and treatment of one case ENKTL patients with EBV negative were analyzed retrospectively.@*RESULTS@#A 46-year-old woman diagnosed as positive for exosanal NK/T cell lymphoma (EBER@*CONCLUSION@#EBV negative ENKTL is rare in clinic and easy to be misdiagnosed, so it should be distinguished from peripheral T cell lymphoma. This case was treated with EBV positive ENKTL regimen, with good short-term efficacy.

Controversies on the values for health instruments of Chinese medicine / 中国结合医学杂志

Currently, there are increasing debates on the necessity of health instruments in Chinese medicine (CM) emerging in China. This study aims to reevaluate its status and values. Analyzing the causes, limits, advantages, and properties characters of health instruments in CM, it is found that weak fundamental research, incomplete self-awareness, and complicated social factors are the primary causes of debates. A comprehensive analysis showed health instruments in CM have health evaluation benefits to people from a dominant Chinese culture, meet the requirements of cultural background, and bring long-term value to Chinese instrument researches. However, its values and status should be treated differently depending on various subtypes. Although little theoretical and practical evidences proved that patients-reported health instruments in CM should be proposed independently, the doctors- and nurses-reported questionnaires are necessary. With this in mind, the study group proposes the 'Chinese cultural instruments (CCIs)' and 'health-related CCIs'. The latter one aims to evaluate the health status of people in a dominant Chinese culture. The CCIs theory represents Chinese instrument researches on a larger regional and higher level, and resolves the debates on instruments between CM and Western medicine in China. Health instruments in CM bring more scientific and social benefits for Chinese instrument researches. However, it does not include cultural demands, and lacks scientific significance. CCIs have all its virtues, and add solutions to the latter's theory bottleneck and scientific debates, thus bringing increased benefits to clinical assessment in complementary and alternative medicine researches.

Culturing primary hippocampal neurons of neonatal mouse and morphologic observation / 重庆医学

Objective To discuss a optimal culture method of primary hippocampal neurons and a more suitable method of mor-phological observation ,and provide basis to the study of synapse in Alzheimer′s Disease .Methods Postnatal 0 -1 days (P0 -1 ) C57BL/6J mice were decollated and bilateral hippocampus were separated .Low level concentration of trypsin and mechanical disso-ciation were adopted .And culture medium without serum was used to culture neurons .After 17 days culturing ,transfected neurons with Green Fluorescent Protein(GFP) by calcium phosphate precipitation ,and then observed neurons and spines by fluorescence mi-croscope .Results The neurons looked good and healthy by using this method .And the axons ,dendrites and spines which were typ-ical structure of neurons were observed clearly after transfected with GFP .Conclusion The cultured hippocampal neurons look good by this method .And the morphological characteristics of neurons and spines are observed much more clearly after transfected GFP by calcium phosphate precipitation .

Construction and immunogenicity of a recombinant adenovirus co-expressing the E2 protein of classical swine fever virus and the porcine interleukin 2 in rabbits / 病毒学报

To construct a recombinant adenovirus co-expressing the E2 protein of classical swine fever virus (CSFV) and the porcine interleukin 2 (pIL-2), the CSFV E2 gene and pIL-2 gene were amplified respectively from the plasmids pMD19-T-E2 and pMD19-T-pIL-2 by PCR. E2-pIL-2 fusion gene was obtained by using 5 consecutive glycine codons as a linker and cloned into the adenoviral shuttle plasmid AdTrack. The AdTrack-E2-pIL-2 was linearized and transformed into E. coli BJ5183 with the backbone plasmid AdEasy1. The resultant recombinant plasmid AdEasy-E2-pIL-2 was transfected into the 293 cells where the recombinant adenovirus rAd-E2-pIL-2 was produced. The immunogenicity of rAd-E2-pIL-2 was evaluated in rabbits. The results of RT-PCR and Western-blotting showed that rAd-E2-pIL-2 could carry and express E2 and pIL-2 proteins. The titer of the rAd-E2-pIL-2 was 10(8.12) PFU/mL. After immunized with rAd-E2pIL-2, The injected rabbits developed a high level of CSFV specific antibodies. Regular fever was not detected in the rAd-E2-pIL-2-immunized rabbits upon challenge with CSFV C stain, and specific lymphoproliferative responses to the CSFV was detected in the lymphocytes from the immunized rabbits. In conclusion, rAd-E2-pIL-2 was constructed successfully and it could be an attractive vaccine candidate against CSFV.