Objective To study the effects of thymus transplantation(TT)combined with CD4--DLI on T cell reconstitution after allogeneic hematopoietic stem cell transplantation(allo-HSCT). Methods BALB/c mice were randomly divided into three groups:hematopoietic stem cell transplantation (HSCT group),hematopoietic stem cell transplantation combined with thymus transplantation(TT group)and hematopoietic stem cell transplanta-tion combined with thymus transplantation plus CD4+ T cell-depleted lymphocyte infusion(CD4--DLI group). On day-1,the mice were treated with the lethal dose of radiotherapy. On day 0,C57BL/6 mice were used as donor for hematopoietic stem cell transplantation. The mice were sacrificed on 5 days,2 weeks,4 weeks and 3 months after transplantation,respectively. The peripheral blood and spleen cells of mice were collected for determinations of T cell surface antigen,T cell receptor,naive T cells and intracellular cytokines. HE staining was used to assess the development of donor thymus. Results TT and CD4--DLI did not impair each other′s effects on T cell reconstitu-tion. TT combined with CD4--DLI increased the number of T cell reconstruction. CD4--DLI promoted the effect of TT on enlargement naive CD4+and CD8+T cell pool. Combination of TT and CD4--DLI enhanced the cytokine pro-duction of T cells. Conclusion TT combined with CD4--DLI had no side effects on TCR repertoire and thymus. Conclusion TT combined with CD4--DLI can enhance the reconstitution of T cell number and function via thymus dependent and thymus independent mechanism.

Background and purpose:For stageⅠ non-small cell lung cancer (NSCLC), video-assisted thoracic segmentectomy is given much attention to by thoracic surgeon because of the less tissue damages. However, video-assisted thoracic lobectomy is still considered as the standard treatment in the world. Therefore, this study was to evaluate the clinical effect after video-assisted thoracic segmentectomy and lobectomy in patients with stageⅠ NSCLC in order to provide reference for clinical application.Methods:The comparative studies on video-assisted thoracic segmentectomy and lobectomy treating stage I NSCLC were retrieved from PubMed, Web of Science, EMBASE, the Cochrane Library, CNKI, CBM, VIP, and Wanfang Data. All data were acquired until July 2015. Literature screening according to data extraction and quality assessment was completed by two reviewers independently. Meta-analysis was conducted by RevMan 5.3 software which was offered by Cochrane network.Results:A total of 11 articles involving 1 677 patients were ifnally included. The results of meta-analysis indicated that: for stageⅠ NSCLC, compared with video-assisted thoracic lobectomy, the effect of video-assisted thoracic segmentectomy was alike in total mortality (OR=0.77, 95%CI: 0.48 to 1.21,P=0.25), 5-year mortality (OR=0.77, 95%CI: 0.52 to 1.14,P=0.19) and systemic complications (OR=0.76, 95%CI: 0.53 to 1.09,P=0.13), but could reduce blood loss [difference in means (MD)=-41.16, 95%CI: -59.46 to -22.86,P<0.000 1], chest tube duration (MD=-0.29, 95%CI: -0.49 to -0.09,P=0.005) and the length of hospital stay (MD=-0.74, 95%CI: -1.44 to -0.05,P=0.04).Conclusion:Compared with video-assisted thoracic lobectomy, video-assisted thoracic segmentectomy can signiifcantly reduce blood loss, chest tube duration and length of hospital stay. However, the two kinds of operation methods achieved the same effects on the total mortality, 5-year mortality and systemic complications. Thoracoscopic segmentectomy may be an alternative to thoracic lobectomy.

OBJECTIVEThe aim of this study was to establish a standardized protocol for detection of ALK protein expression and gene fusion in cytologic specimens.

METHODSLung adenocarcinoma cytologic specimens were collected from seven hospitals in Beijing city. A detection protocol for ALK protein expression and gene fusion was designed according to the results of comparative experiment. Ventana immunohistochemical (IHC) ALK(D5F3) detecting ALK protein expression was performed in 203 prepared formalin-fixed paraffin-embedded (FFPE) cell blocks. ALK gene fusion in 98 EGFR gene wild type cytologic specimens and in 4 bronchoalveolar lavage fluid (BL) samples was detected by quantitative reverse transcription polymerase chain reaction (qRT-PCR). ALK gene fusion in the Ventana IHC ALK (D5F3) positive samples was further tested by fluorescence in situ hybridization (FISH). Six patients with ALK IHC-positive result were followed up to analyze the responses of crizotinib therapy. Comparative experiments: (1) Comparison of the results of 4% neutral buffered formalin fixed for different time (24 h, 48 h, 72 h) on the Ventana IHC ALK (D5F3) staining was conducted in two cases of IHC ALK positive FFPE cell blocks; (2) Comparing qRT-PCR results for ALK fusion in samples from FFPE cell blocks and cytospin prepared slides in 10 cases of lung adenocarcinoma cytologic specimens.

RESULTSAmong the specimens examined using the standardized protocol recommended by this study, 229 cases of cytologic specimens met the diagnostic criteria of lung adenocarcinoma. Among them, 207 cases obtained ALK gene test results (by at least one method), with an ALK test ratio of 90.4% (207/229). FFPE cell blocks were successfully prepared in 203 cases, Ventana IHC ALK (D5F3) were successfully performed in all the 203 FFPE cell blocks (100%), and the ALK protein positive detection rate was 10.3% (21/203). ALK fusion was tested in 98 FFPE cytologic samples of EGFR wild types by qRT-PCR, and 96 out of 98 (97.96%) cytologic samples were successfully performed.18 out of 19 IHC ALK-positive cases were verified to be of ALK fusion status by qRT-PCR. The concordance rate was 94.7% (Kappa=0.967, P<0.001) between Ventana IHC ALK (D5F3) and qRT-PCR, and the sensitivity of the Ventana IHC ALK (D5F3) assay compared with qRT-PCR was 100% and the specificity was 98.7%. FISH assay was used to verify the positive cases detected by Ventana IHC ALK (D5F3) staining. Two cases of low tumor cell content FFPE samples obtained indefinite results by FISH test. The six patients with positive ALK protein expression received crizotinib therapy, and 5 paitents got treated effectively. For two ALK IHC positive cases, which were 4% neutral buffered formalin fixed for 72 h, the result of Ventana IHC ALK (D5F3) staining became weakened obviously and uneven. In 10 cases of samples, total RNA was extracted from FFPE cytologic sections and cytospin prepared slides, and the results of qRT-PCR test and ALK gene fusion showed good concordance.

CONCLUSIONSThe standardized protocol recommended in this study expands the detection types and quantity of cytologic specimens for ALK protein expression and gene fusion and increased the detection rate. Ventana IHC ALK (D5F3) is a reliable method for detecting ALK protein expression in FFPE cell blocks. The pathologic quality control procedure prior to Ventana IHC ALK (D5F3) is crucial for the accuracy of testing the ALK gene status. When FFPE cell blocks could not be prepared or prepared unsuccessfully from the cytologic specimens, qRT-PCR may be an alternative option for the detection of ALK gene fusion.

Adenocarcinoma ; drug therapy ; enzymology ; genetics ; pathology ; Alkaline Phosphatase ; genetics ; metabolism ; Gene Fusion ; Genes, erbB-1 ; Humans ; Immunohistochemistry ; In Situ Hybridization, Fluorescence ; Lung Neoplasms ; drug therapy ; enzymology ; genetics ; pathology ; Protein Kinase Inhibitors ; therapeutic use ; Proteomics ; Pyrazoles ; therapeutic use ; Pyridines ; therapeutic use ; Sensitivity and Specificity

OBJECTIVEBenign epilepsy with centro-temporal spikes (BECTs) is a common idiopathic partial epileptic syndrome in childhood, which often affect the pre-school and school-age children and a considerable proportion have comorbidity including lower academic achievement and cognitive impairment. Few studies involved the psychocognitive assessment in such a drug-treatable epileptic syndrome especially in the newly diagnosed and medications-naive group. This study aimed to investigate the cognitive characteristics of children with newly onset BECTs before treatment.

METHODForty-one outpatients with newly diagnosed BECTs who visited the Clinic during the periods from October 2012 to May 2014 before the medications against epilepsy and 41 healthy controls recruited from regular school in Beijing during the period from July 2013 to March 2014, who matched in age and gender underwent battery testing by computerized cognitive testing in epilepsy (CCTE). The BECTs group included 41 children, 20 boys and 21 girls, mean age (8.2 ± 1.7) years, the age of onset of epilepsy 4.5-11.5 years (the age of onset <8 years in 25 cases, ≥ 8 years in 16 cases). The cognitive characteristics and associated factors were analyzed. The primary data including correct answer numbers and reaction times were analyzed by independent sample t-test between the two groups of children with BECTs and healthy controls based on SPSS 18.0 statistical software.

RESULTRaw data of 9 tasks' scores collected from BECTs and healthy control children were continuous variables in accordance with normal distribution. BECTs children performed significantly worse than controls in choice reaction time ((618+158) vs. (524+254) ms), three-dimensional mental rotation (11 ± 10 vs. 18 ± 12) and visual tracing (10 ± 6 vs.15 ± 6), t=2.01, 3.03 and 3.47, P<0.05, <0.01 and <0.001, respectively.While other 6 tasks showed no significant difference between the two groups (P>0.05 for all comparisons). BECTs boys performed significantly worse than girls on simple substraction tasks compared with standard nine score ((4.7 ± 1.5) vs. (5.6 ± 1.2), t=-2.24, P<0.05). Other 8 tasks showed no significant difference between boys and girls (P>0.05 for all comparisons). Other 9 tasks showed no significant differences between the two groups of BECTs children whose age of onset was before 8 years and those who started seizure ≥ 8 years (P all >0.05). The standard nine scores of simple substraction from the three BECTs groups of dominance sides of spikes and waves during NREM showed significant difference (P<0.05). BECTs children with bilateral discharges performed significantly worse than the other two groups dominantly right or left discharges (4.7 ± 1.2 vs. 6.0 ± 1.2 vs. 4.9 ± 1.4, P all <0.05). There was no significant difference between the two groups with right and left side dominance discharges (P>0.05). Other 8 tasks showed no significant differences among the three groups (P>0.05 for all comparisons).

CONCLUSIONAlthough EEG discharges index below 50% during NREM period, while newly diagnosed BECTs children before treatment with medications against epilepsy performed poorer on tasks of choice reaction time, three-dimensional mental rotation, and visual tracing. The two factors of male and bilateral discharges during NREM period correlate with dysfunction of simple subtraction, the mechanism needs further study and the cognitive function of epilepsy children should be evaluated and followed up, in order to provide psychologic baseline data for persistent cognitive disturbance.

Beijing ; Case-Control Studies ; Child ; Child, Preschool ; Cognition ; Cognition Disorders ; diagnosis ; Comorbidity ; Epilepsy, Rolandic ; physiopathology ; Female ; Humans ; Male ; Reaction Time ; Seizures ; physiopathology

Objective The aim of this study was to establish a standardized protocol for detection of ALK protein expression and gene fusion in cytologic specimens. Methods Lung adenocarcinoma cytologic specimens were collected from seven hospitals in Beijing city. A detection protocol for ALK protein expression and gene fusion was designed according to the results of comparative experiment. Ventana immunohistochemical ( IHC) ALK( D5F3) detecting ALK protein expression was performed in 203 prepared
formalin?fixed paraffin?embedded ( FFPE) cell blocks. ALK gene fusion in 98 EGFR gene wild type cytologic specimens and in 4 bronchoalveolar lavage fluid ( BL ) samples was detected by quantitative reverse transcription polymerase chain reaction (qRT?PCR). ALK gene fusion in the Ventana IHC ALK (D5F3) positive samples was further tested by fluorescence in situ hybridization ( FISH) . Six patients with ALK IHC?positive result were followed up to analyze the responses of crizotinib therapy. Comparative experiments:( 1) Comparison of the results of 4% neutral buffered formalin fixed for different time ( 24 h, 48 h, 72 h) on the Ventana IHC ALK (D5F3) staining was conducted in two cases of IHC ALK positive FFPE cell blocks;(2) Comparing qRT?PCR results for ALK fusion in samples from FFPE cell blocks and cytospin prepared slides in 10 cases of lung adenocarcinoma cytologic specimens. Results Among the specimens examined using the standardized protocol recommended by this study, 229 cases of cytologic specimens met the diagnostic criteria of lung adenocarcinoma. Among them, 207 cases obtained ALK gene test results ( by at least one method), with an ALK test ratio of 90.4% (207/229).FFPE cell blocks were successfully prepared in 203 cases, Ventana IHC ALK ( D5F3) were successfully performed in all the 203 FFPE cell blocks ( 100%) , and the ALK protein positive detection rate was 10.3% (21/203). ALK fusion was tested in 98 FFPE cytologic samples of EGFR wild types by qRT?PCR, and 96 out of 98 ( 97. 96%) cytologic samples were successfully performed.18 out of 19 IHC ALK?positive cases were verified to be of ALK fusion status by qRT?PCR. The concordance rate was 94.7% ( Kappa=0.967, P<0.001) between Ventana IHC ALK( D5F3) and qRT?PCR, and the sensitivity of the Ventana IHC ALK ( D5F3) assay compared with qRT?PCR was 100%and the specificity was 98. 7%. FISH assay was used to verify the positive cases detected by Ventana IHC ALK(D5F3) staining. Two cases of low tumor cell content FFPE samples obtained indefinite results by FISH test. The six patients with positive ALK protein expression received crizotinib therapy, and 5 paitents got treated effectively. For two ALK IHC positive cases, which were 4% neutral buffered formalin fixed for 72 h, the result of Ventana IHC ALK(D5F3) staining became weakened obviously and uneven. In 10 cases of samples, total RNA was extracted from FFPE cytologic sections and cytospin prepared slides, and the results of qRT?PCR test and ALK gene fusion showed good concordance. Conclusions The standardized protocol recommended in this study expands the detection types and quantity of cytologic specimens for ALK protein expression and gene fusion and increased the detection rate. Ventana IHC ALK( D5F3) is a reliable method for detecting ALK protein expression in FFPE cell blocks. The pathologic quality control procedure prior to Ventana IHC ALK( D5F3) is crucial for the accuracy of testing the ALK gene status. When FFPE cell blocks could not be prepared or prepared unsuccessfully from the cytologic specimens, qRT?PCR may be an alternative option for the detection of ALK gene fusion.

Objective The aim of this study was to establish a standardized protocol for detection of ALK protein expression and gene fusion in cytologic specimens. Methods Lung adenocarcinoma cytologic specimens were collected from seven hospitals in Beijing city. A detection protocol for ALK protein expression and gene fusion was designed according to the results of comparative experiment. Ventana immunohistochemical ( IHC) ALK( D5F3) detecting ALK protein expression was performed in 203 prepared
formalin?fixed paraffin?embedded ( FFPE) cell blocks. ALK gene fusion in 98 EGFR gene wild type cytologic specimens and in 4 bronchoalveolar lavage fluid ( BL ) samples was detected by quantitative reverse transcription polymerase chain reaction (qRT?PCR). ALK gene fusion in the Ventana IHC ALK (D5F3) positive samples was further tested by fluorescence in situ hybridization ( FISH) . Six patients with ALK IHC?positive result were followed up to analyze the responses of crizotinib therapy. Comparative experiments:( 1) Comparison of the results of 4% neutral buffered formalin fixed for different time ( 24 h, 48 h, 72 h) on the Ventana IHC ALK (D5F3) staining was conducted in two cases of IHC ALK positive FFPE cell blocks;(2) Comparing qRT?PCR results for ALK fusion in samples from FFPE cell blocks and cytospin prepared slides in 10 cases of lung adenocarcinoma cytologic specimens. Results Among the specimens examined using the standardized protocol recommended by this study, 229 cases of cytologic specimens met the diagnostic criteria of lung adenocarcinoma. Among them, 207 cases obtained ALK gene test results ( by at least one method), with an ALK test ratio of 90.4% (207/229).FFPE cell blocks were successfully prepared in 203 cases, Ventana IHC ALK ( D5F3) were successfully performed in all the 203 FFPE cell blocks ( 100%) , and the ALK protein positive detection rate was 10.3% (21/203). ALK fusion was tested in 98 FFPE cytologic samples of EGFR wild types by qRT?PCR, and 96 out of 98 ( 97. 96%) cytologic samples were successfully performed.18 out of 19 IHC ALK?positive cases were verified to be of ALK fusion status by qRT?PCR. The concordance rate was 94.7% ( Kappa=0.967, P<0.001) between Ventana IHC ALK( D5F3) and qRT?PCR, and the sensitivity of the Ventana IHC ALK ( D5F3) assay compared with qRT?PCR was 100%and the specificity was 98. 7%. FISH assay was used to verify the positive cases detected by Ventana IHC ALK(D5F3) staining. Two cases of low tumor cell content FFPE samples obtained indefinite results by FISH test. The six patients with positive ALK protein expression received crizotinib therapy, and 5 paitents got treated effectively. For two ALK IHC positive cases, which were 4% neutral buffered formalin fixed for 72 h, the result of Ventana IHC ALK(D5F3) staining became weakened obviously and uneven. In 10 cases of samples, total RNA was extracted from FFPE cytologic sections and cytospin prepared slides, and the results of qRT?PCR test and ALK gene fusion showed good concordance. Conclusions The standardized protocol recommended in this study expands the detection types and quantity of cytologic specimens for ALK protein expression and gene fusion and increased the detection rate. Ventana IHC ALK( D5F3) is a reliable method for detecting ALK protein expression in FFPE cell blocks. The pathologic quality control procedure prior to Ventana IHC ALK( D5F3) is crucial for the accuracy of testing the ALK gene status. When FFPE cell blocks could not be prepared or prepared unsuccessfully from the cytologic specimens, qRT?PCR may be an alternative option for the detection of ALK gene fusion.

OBJECTIVEThe aim of this study was to establish a standard protocol for detection of EGFR mutations in cytologic specimens.

METHODS287 cytologic samples were collected from the patients who were suspected of having lung cancer at six hospitals in Beijing. A detection protocol for EGFR mutations was designed. Two comparative experiments were carried out for the coincidence in EGFR mutation rates between direct sequencing (Seq) and amplification refractory mutation system (ARMS) methods, and between 40 matched cytologic samples with formaldehyde-fixed paraffin embedded (FFPE) cytologic blocks and cytospin slides.

RESULTSTumor cells were found in 236 out of 287 cases (82.2%, 236/287) . Among them, there were 31 cases (13.1%, 31/236) of low tumor cell content samples and 205 cases (86.9%, 205/236) of high tumor cell content samples. 180 cases in the high tumor cell content samples (87.8%, 180/205) were diagnosed to be consistent with NSCLC. 25 out of 194 cases were ruled out or indefinite to be diagnosed as NSCLC by immunohistochemistry. By direct sequencing, the mutation rate of EGFR was 27.8% (50/180) in NSCLC samples and 28.2% (50/177) in adenocarcinoma samples (high tumor content samples) . By ARMS, the mutation rate of EGFR was 45.6% (82/180) in NSCLC samples and 46.3% (82/177) in adenocarcinoma samples (high tumor content samples). The EGFR mutation rate in low tumor content samples was 38.7% (12/31) , there was no significant difference in EGFR mutation rates between the groups of low tumor cell content samples and high tumor cell content samples (P = 0.12). The concordance rate of EGFR mutation rates was 100% between scraping tumor cells from slides samples and from FFEP blocks in the 40 matched samples. Forty-eight out of 180 definitive NSCLC patients received Gefitinib therapy. The FPS was 12 months in the gefitinib-treated ARMS⁺ group and 2 months in the ARMS⁻ group (P < 0.001), and the OS was 19 months in the gefitinib-treated ARMS⁺ group and 7 months in the ARMS⁻ group (P = 0.003), but no significant differences were found in the efficacy (PFS and OS) of Gefitinib between Seq⁺ and Seq⁻ groups (P = 0.227, P = 0.510, respectively), and Seq⁺/ARMS⁺ and Seq⁻/ARMS⁺ groups (P = 0.354, P = 0.334, respectively).

CONCLUSIONSThe detection protocol for EGFR mutations in cytological specimens introduced in this study is tested to be reliable and feasible. Pathological evaluation and immunohistochemistry are important in the detection procedure of EGFR mutations in cytologic specimens. High sensitivity methods should be selected for detection of EGFR mutations in cytologic samples.

Adenocarcinoma ; metabolism ; Carcinoma, Non-Small-Cell Lung ; metabolism ; Humans ; Lung Neoplasms ; diagnosis ; epidemiology ; metabolism ; Mutation ; Mutation Rate ; Polymerase Chain Reaction ; Receptor, Epidermal Growth Factor ; genetics ; metabolism

Objective To observe the effects of comprehensive warming measures on children ’ s anesthesia recovery , in order to prevent the adverse effects of the anesthesia period .Methods In this prospective study , 60 children from July 2012 to July 2013 entering PACU were randomly divided into the experimental group and the control group , with 30 cases each . The experimental group was based on comprehensive warming measures .The control group was based on conventional warming method .The residence time in PACU, mean arterial pressure, heart rate, oxyhemoglobin saturation, body temperature changes and the complication rate in the anesthesia recovery period were compared between groups .Results The PACU residence time of the experimental group was (57 ±15)min, which was significantly lower than (96 ±18)min of the control group (t=6.342,P<0.05).The rate of chill during anesthesia recovery period was 10.0%in the experimental group, which was significantly lower than 33.3% in the control group (χ2 =7.134,P<0.05). The incidence of hypoxemia , agitation and laryngospasm had no significant differences between the two groups (P >0.05 ).Conclusions The comprehensive warming measures can reduce the PACU residence time , stabilize the hemodynamic parameters , reduce the incidence of chill and improve the safety and nursing quality during children ’ s anesthesia recovery period .

OBJECTIVETo investigate whether intracerebral hemorrhage (ICH) can promote neurogenesis in the dentate gyrus (DG) of rat hippocampus.

METHODSWestern blot analysis, immunohistochemical staining, and immunofluorescent double labeling combined with confocal microscope were used to detect neurogenesis in the DG of the hippocampus in rats after ICH.

RESULTSThe expression of DCX protein in the ipsilateral DG of the hippocampus was enhanced in the rats 1 day after ICH (0.202∓0.062) as compared with that in normal rats (0.127∓0.088), reaching the peak level at 14 days (0.771∓0.108, P<0.01) and beginning to decrease at 28 days (0.582∓0.121, P<0.01). Meanwhile, DCX-positive cells and BrdU-positive cells, and DCX/BrdU double-labeled cells were detected in the DG of the hippocampus. Compared with those in the control group, BrdU/NeuN double-labeled cells were markedly increased in the granular cell layer of the DG at 28 days after ICH (1.808∓1.020 vs 5.654∓1.671, P<0.01).

CONCLUSIONICH can promote neurogenesis in the DG of rat hippocampus.

Animals ; Antigens, Nuclear ; metabolism ; Bromodeoxyuridine ; metabolism ; Cerebral Hemorrhage ; metabolism ; pathology ; physiopathology ; Dentate Gyrus ; metabolism ; physiopathology ; Hippocampus ; metabolism ; physiopathology ; Microtubule-Associated Proteins ; metabolism ; Nerve Tissue Proteins ; metabolism ; Neurogenesis ; physiology ; Neurons ; metabolism ; physiology ; Neuropeptides ; metabolism ; Rats ; Rats, Sprague-Dawley

Objective To investigate whether intracerebral hemorrhage (ICH) can promote neurogenesis in the dentate gyrus (DG) of rat hippocampus. Methods Western blot analysis, immunohistochemical staining, and immunofluorescent double labeling combined with confocal microscope were used to detect neurogenesis in the DG of the hippocampus in rats after ICH. Results The expression of DCX protein in the ipsilateral DG of the hippocampus was enhanced in the rats 1 day after ICH (0.202 ± 0.062) as compared with that in normal rats (0.127 ± 0.088), reaching the peak level at 14 days (0.771 ± 0.108, P<0.01) and beginning to decrease at 28 days (0.582±0.121, P<0.01). Meanwhile, DCX-positive cells and BrdU-positive cells, and DCX/BrdU double-labeled cells were detected in the DG of the hippocampus. Compared with those in the control group, BrdU/NeuN double-labeled cells were markedly increased in the granular cell layer of the DG at 28 days after ICH (1.808 ± 1.020 vs 5.654 ± 1.671, P<0.01). Conclusion ICH can promote neurogenesis in the DG of rat hippocampus.