Advances in molecular mechanisms of organelle interaction and their role in disease developmen / 中国药科大学学报

@#Organelles have their special functions, they interact with each other and coordinate a series of important physiological functions at the same time. Organelle interaction occurs at membrane contact sites(MCSs), where the membranous organelle endoplasmic reticulum is the core, and specific tethered proteins at the membrane contact site bind to the organelle membrane and various protein complexes work together to perform specific functions, such as lipid transport, Ca2+ transfer, etc. This review studies on the structure and function of membrane contact sites and their key roles in organelle interactions, focusing on the connection between the endoplasmic reticulum and plasma membrane, mitochondria and Golgi, as well as the association between the key proteins at membrane contact sites and the occurrence and development of various diseases.

Establishment and application of flow cytometry in detecting human cells in human/goat chimera models / 中华检验医学杂志

Objective To establish the methodology of flow cytometry for detecting human cells in human/goat chimerisra.Methods Human hemopoietic stem/progenitor cells (CD+34 cells) or MIG-tranadueed-GFP CD+34 cells were transplanted into the peritoneal cavity of fetal goats in utero to obtain human/goat chimera modeL The peripheral blood cells from the chimeras were labeled with multiple mouse anti-human antibodies and the monoelonal antibodies that were specific for human but had not or only minimal cross-reaction with goat were screened as the primary antibodies for routine analysis in flow cytometry.Human cord blood was proportionally (25% ,50% ,75%,100%) added into the blood of the untransplanted goats and the cells were labeled with CD+34 monoclonal antibody.The region and size of the "gate" were chosen based on to the distribution of CD+34 cells or human cord blood.One human/goat chimera marked with GFP (MIG goat) was sacrificed and the substantial liver cells from its perfused liver were analyzed for the GFP+cells percentage and DNA contents by flow cytometry.Results CD7,CD15,CD38,CD45CD20CD34CD14and GPA monoclonal antibodies were chosen as the primary antibodies in rou tine detection by flow cytometry.The size and area of the "gate" were also defined.29.1% (29100/100 000 ) of the substantial liver cells from the MIG goat expressed GFP.DNA content analysis showed that the GFP+ cells obtained from the liver of MIG goat mainly manifested two peaks that were correspond to those of human.Conclusions Flow cytometry is rapid,simple and effective for the investigation of differentiation,homing and biological characteristics of stem cells in vivo.The selections of suitable surface antibodies and the "gate" are very important for detecting human cells accurately in the human/goat chimerism.

An Expedient Reliable Double Fluorescent Reporter System for φC31 Integrase Function Evaluation / 生物化学与生物物理进展

A reporter system for φC31 integrase was developed in NIH3T3 cells. The reporter plasmid coding green fluorescent protein (GFP) coupled with red fluorescent protein (RFP) was eo-transfected with the plasmid coding φC31 integrase, to show the activity of integrase in the cells. Fluorescence activated cell sorter (FACS) was used to measure the proportion of the cells containing red and green fluorescence. The increment of green cells was positively related to the increase in the transfection with plasmid coding φC31 integrase. Approximately 90% of green cells were observed under a ratio of [plasmid-φC31-integrase]/[reporter plasmid] at 10 : 1. This suggests that the φC31 integrase reporter system provides a probe for the function of φC31 integrase in cells.

An Expedient Reliable Double Fluorescent Reporter System for ?C31 Integrase Function Evaluation / 生物化学与生物物理进展

A reporter system for ?C31 integrase was developed in NIH3T3 cells.The reporter plasmid coding green fluorescent protein(GFP) coupled with red fluorescent protein(RFP) was co-transfected with the plasmid coding ?C31 integrase, to show the activity of integrase in the cells.Fluorescence activated cell sorter(FACS) was used to measure the proportion of the cells containing red and green fluorescence.The increment of green cells was positively related to the increase in the transfection with plasmid coding ?C31 integrase.Approximately 90% of green cells were observed under a ratio of plasmid-?C31-integrase/reporter plasmid at 10∶1.This suggests that the ?C31 integrase reporter system provides a probe for the function of ?C31 integrase in cells.

Detection of human/goat xenogeneic models by interphase fluorescence in situ hybridization / 中华医学遗传学杂志

<p><b>OBJECTIVE</b>To establish a high sensitive and specific method of interphase fluorescence in situ hybridization (IFISH) to detect the low-frequency human cells in human/goat xenogeneic models.</p><p><b>METHODS</b>Human-specific Y-chromosome satellite DNA CEPY and 17-chromosome satellite DNA p17H8 were used as probes for IFISH. The peripheral blood samples from 2 goats transplanted with human male hematopoietic stem cells (HSC), 1 normal negative goat and 1 normal man were analyzed. The actual FISH efficiency was confirmed by serial dilutions (1/100, 1/500 and 1/1000) of the cell mixture of normal man and normal negative goat. A set of signal scoring criteria was determined to guarantee the stability and reliability of the method.</p><p><b>RESULTS</b>Positive cell (human cell) frequencies were consistent with the established frequencies for the human/goat cell mixture. The average frequencies of positive cells were 98.60% (CEPY) and 100% (p17H8) for normal man, 0 for normal negative goat, 0.23% (CEPY) and 0.11% (p17H8) for human/goat xenogeneic models. The results demonstrated that low-frequency human cells (male cells confirmed by Y-chromosome probe) existed in human/goat xenogeneic models.</p><p><b>CONCLUSION</b>The IFISH developed in this study is of high sensitivity and specificity and can identify the actual frequency of human cells, which offers a direct, sensitive and specific approach to the detection of low-frequency human cells in human/goat xenogeneic models.</p>

Analysis of the status of mouse fetal transgenic cells existed in the maternal peripheral blood / 中华检验医学杂志

Objective To study the status of mouse fetal transgenic cells existed in the maternal blood in peri-natal period and provide the information for gene diagnosis using fetal cells circulated in the maternal blood.Methods Mating the female normal mice with male transgenic mice integrated with GFP reporter gene driven by ?-globin promoter and HS2-HS3 elements of LCR. Around the offspring were born, maternal blood was collected and GFP expression level was determined with FACS analysis. Meanwhile, DNA extracted from the tails of the mothers and their offspring as well as the maternal blood were analyzed by PCR.Results GFP positive fetal cells were not found in maternal blood before offspring were born. 1-3 weeks later, GFP positive fetal cells were detected and this population in maternal peripheral blood reached the peak. 4-5 weeks later, they disappeared gradually. PCR results showed no GFP positive band in the mothers. However, a positive fragment of GFP gene was amplified in maternal blood samples after their offspring were born. This result was in accordance with FACS analysis.Conclusion Transgene cells may be useful markers for the study of status of fetal cells existed in the maternal circulating blood. The results would be beneficial for the gene diagnosis using maternal blood as an alternative resource of fetal cells.

Molecular identification of human/goat xenogeneic model / 中华血液学杂志

<p><b>OBJECTIVE</b>To identify the human hematopoietic stem cells from the human/goat xenogeneic model with molecular techniques.</p><p><b>METHODS</b>DNA and total RNA were extracted from 11 transplanted goat peripheral blood cells. Human CD(34), GPA and SRY genes were amplified with PCR in these samples, and CD(34), GPA mRNA transcripts were detected using RT-PCR in 5 and 6 goat peripheral blood cells, respectively. Southern blot analysis was performed in 8 goat DNAs to detect the human specific alpha-satellite sequence. Meanwhile FISH was also performed to detect the human cells in goat blood with a probe of human Y chromosome.</p><p><b>RESULTS</b>Human CD(34) and GPA genes could be detected with PCR in all the 11 goats, and SRY gene did in 5 goats transplanted with hematopoietic stem cells derived from male human babies. Southern blot showed that human specific alpha-satellite sequence was present in 8 goats. By RT-PCR, human CD(34) mRNA was detected in 5 experimental goats, GPA mRNA was found in the other 6 experimental goats and FISH assay showed that some peripheral blood cells of the human/goat xenogeneic model were positive.</p><p><b>CONCLUSION</b>Existence of human cells in the recipient goats was identified by molecular detection, which was feasible for the examination of human/goat xenogeneic models.</p>

Chest Radiographic Features of Acquired Immunological Deficiency Syndrome(AIDS) After Renal Transplantation / 实用放射学杂志

Objective　To investigate radiographic features of pneumocystis carinii pneumonia(PCP)or Kaposi sarcoma(KS),which were the most common complications of AIDS after renal transplantation.Methods　Radiographic diagnosis and differential diagnosis of 2 cases with PCP and KS comfirmed by pathology were discussed by analysing the chest X-ray findings combined with revieuw of reference literature.Results　The two cases with anti-HIV antibody all had mediasinal lymphopathy.In the case of KS,the omental tuber or tubercle shadows and pleural effusion were found on both pulmonary fields.Conclusion　No characteristic chest radiographic meanifestations in AIDS,the definitive diagnosis depends on clinical laboratory examination and pathological examination.

Cost-effectiveness Analysis of Risperidon and Clozapine in the Treatment of Schizophrenia / 中国药房

0.05).There was a littel difference in the costs for an identical effect between two drugs,the cost of Risperidon being lower.CONCLUSION:The total effective rates and costs of Risperidon and Clozapine were almost equal in the treatment of schizophrenia.