Abstract: Objective To compare the sensitivity and accuracy of autofluorescence bronchoscope (AFB) and white light bronchoscope (WLB) in airway examination for patients with central type lung cancer. Methods From September 2009 to May 2010, 46 patients including 36 males and 10 females with an average age of 62.1 years underwent both AFB and WLB procedures in People’s Hospital of Peking University. Among them, 35 were preliminary diagnostic cases and 11 were postoperative surveillance cases. Local anaesthesia of glottis and airway, and general anaesthesia with continuous intravenous drugs were given before electric bronchoscope was adopted. All patients underwent WLB examination followed by AFB procedure. All suspicious abnormal visual findings were recorded for biopsy and pathological examination. Results All procedures were carried out safely without death or severe complications. We performed bronchoscopy 48 times for all 46 patients and 159 tissues of various sites were taken out for biopsy and pathologic examination which showed 64 malignancies and 95 none malignancies. In 64 malignancies, AFB found all but WLB missed 15 with a missed diagnosis rate of 23.4%. Thirtysix times of examination were performed for the 35 preliminary diagnostic cases and 56 sites of malignancy were found. AFB found all, while WLB missed 12, and 6 sites of malignancy found by AFB were larger in size than those found by WLB. AFB detected 3 cases of multisite malignancy, but WLB missed these diagnoses. The results of AFB and WLB were the same for 26 patients. Twelve times of bronchoscopy were performed for the 11 postoperative surveillance cases and 8 sites of malignancy were found. AFB found them all while WLB missed 3 which were two recurrent cases during the early period after lung cancer surgery. The sensitivity of AFB and WLB was 100.0 % and 76.6%(Plt;0.05) respectively, and the negative predictive value of AFB and WLB was 100.0% and 84.5%(P=0.002) respectively. Conclusion AFB has a better sensitivity and negative predictive value than WLB in detecting mucous canceration lesions in central type lung cancer, and is more accurate in assessment of tumor margins, more sensitive in finding multiple lesions in airway and detecting early cancer recurrence in postoperative surveillance patients.
Objective To insure early detection and hence efficient prevention of allograft rejection in transplanted heart, investigate possible applications of NAD(P)H fluorescence components analysis at the level of living cardiac cells to propose new approaches for diagnosis of rejection. Methods NAD(P)H was studied for noninvasive fluorescent probing of the mitochondrial function. Human cardiomyocyte were isolated from one additional endomyocardial biopsy (EMB) of 14 pediatric patients with heart ransplantation. Rat cardiomyocyte (n=5, 13-14 week old) were also isolated by the same approach for human myocytes. Autofluorescence(AF) was recorded in living cardiomyocytes following excitation with 375 nm UVlight and detection by spectrallyresolved time correlated single photon counting (TCSPC), based on the simultaneous measurement of the fluorescence spectra and lifetimes. Rat cardiac cells were divided into four groups: normoxic condition, normoxia with Rotenone, ischemic condition and ischemia with Rotenone. Comparison of cardiomyocyte AF between human and rat; compared kinetics of rat cardiomyocytes AF in normoxic conditions to ischemiamimicking ones, induced at physiological temperatures by reducing cell pH and oxygen content; comparison of cardiomyocyte AF dynamic changes in transplanted pediatric patients presenting either no rejection (R0) or mild rejection (R1). Results We have achieved appropriate isolation of living cardiomyocytes from human biopsies, as well as from rat cardiac tissues and determined their AF. At least a 3-exponential decay with 0.5-0.7ns, 1.9-2.4 ns and 9.0-15.0 ns lifetime pools is necessary to describe human cardiomyocyte AF within 420560 nm spectral range. Rat cardiomyocyte steadystate AF in ischemiamimicking condition was significantly increased when compared normoxic ones (Plt;0.05); application of Rotenone induced a significant increase in AF intensity in ischemic and normoxic condition, however no significant difference between the two groups (Plt;0.05).Human cardiomyocyte AF was found significantly lower in comparison to experimental rat model in the same condition(Plt;0.05). A correlation between changes in steadystate NAD(P)H fluorescence and rejection grades was found when comparison of R1 to R0. R1 showed significantly increased fluorescence intensity (Plt;0.05), without change in the spectra shape, results can be comparable to the effect of ischemiamimic conditions. Conclusion Our studies clearly demonstrated that spectrallyresolved fluorescence spectral analysis coupled to fluorescence lifetime are high sensitive approaches to examine mitochondrial metabolic oxidative state directly in living human cardiomyocytes with good reproducibility. Human cardiomyocytes are more metabolically active than the rat ones, while this activity (and thus ATP production) seems lowered during rejection process. In perspective, the advantage of this method is the possibility of its combination to multiphoton confocal microscopy, which can result in the adaptation of this approach directly to tissue biopsy, as well as in vivo directly via cardiac catheterization without the necessity of cell isolation. This approach provides promising new tool for clinical diagnosis and treatment of allograft rejection, and will enhance our knowledge about cardiomyocyte oxidative metabolism and/or its dysfunction at a cellular level.
Objective To observe preserving effect on myocytes in porcine aortic valve replacement with minimal extracorporeal circulation (MECC). Methods 7 pigs were collected as experimental animals and undertook aortic valve replacement with MECC. Morphological and immunofluorescence intensity changes of right atrial and left ventricular tissues were observed. Results HE staining showed that there were not significant changes and edema or injury of myocytes of right atriums and left ventricles between preoperation and postoperation. Immunofluorescence staining showed complement C3b/c in right atrial myocardial tissues after the operation were a little ber, and innate antibody IgG were a little ber in left ventricular myocardial tissues but similarly weak in right atrial myocardial tissues pre- and post-operation. There was not significant changes in HSPG staining in pre-and post-operative right atrial myocardial tissues, but HSPG were obviously weaker in left ventricular myocardial tissues after the operation. Conclusion MECC is effective on support of porcine aorta valve replacement.
Objective To investigate the method of constructing a tissue engineered epidermis with human epidermal cells and polycarbonate membrane, and to establ ish a tissue engineered epidermis with barrier function which is intended to be the replacing model in vitro of skin irritation test. Methods The tissue engineered epidermis was constructed by using polycarbonate membrane as scaffold and stratified differentiated epidermis derived from human keratinocytes. The tissue engineered epidermis was cultured on an inert polycarbonate filter at the air-liquid interface. After 13 days of culture, the composition and structure of tissue engineered epidermis were observed by HE staining, immunofluorescence staining of keratin 10 (K10) amp; K13, K14, laminin,involucrin, and filaggrin, and transmission electronic microscope. The half maximal inhibitory concentration of a substance (IC50) of SDS was determined in the penetration test of tissue engineered epidermis cultured in the absence (control group) or the presence (experimental group) of l i pid supplement for 18 hours. Results The constructed epidermis was similar to normalepidermis, which was consisted of a proliferating basal layer, differentiated spinous layer, granular layer, and stratum corneum. The IC50 values of tissue engineered epidermis cultured in the control group and experimental group were 0.072% (2.36 mmol/L) and 0.183% (6.00 mmol/L), respectively. Conclusion The tissue engineered epidermis constructed on polycarbonate membrane has normal composition and structure and barrier function corresponding to the normal epidermis.
Objective Using chemically extracted acellular methods to treat extracranial section of the canine whole facial nerve, to evaluated its effects on nerve structure and the removal extent of Schwann cells and myel in. Methods Twenty whole facial nerves were exposed from 10 canines [weighing (18 ± 3) kg]. The extracranial trunk of canine facial nerve and its branches (temporal branch, zygomatic branch, buccal branch, marginal mandibular branch, and cervical branch) were dissected under l ight microscope. Twenty facial nerves were divided into the experimental group (n=12) and control group (n=8) randomly. In experimental group, the nerve was extracted with the 3%TritonX-100 and 4% sodium deoxycholate. In control group, the nerve was not extracted. HE staining and immunofluorescence histological stainings for Hoechst33258, P75, Zero, and Laminin were performed. Results After histological staining, it was found that myel in and Schwann cells were removed from the facial nerve while the basal lamina tube remained intact. The whole canine facial nerves (one nerve trunk and multiple nerve branches) had the similar result. Conclusion The canine whole facial nerve has natural structure (one nerve trunk and multiple nerve branches) by extracted with chemically extracted acellular methods, so it is an available graft for repairing the defect of the whole facial nerve.
Objective To analyze the variation of intestinal microflora in patients with colorectal cancer by SYBR GreenⅠreal-time fluorescence quantitative PCR and reveal the role and significance of intestinal microflora in the colorectal cancer-associated molecular pathogenesis. Methods A set of 16S rRNA gene group of species-specific primers for Bifidobacterium spp., Lactobacillus group, Escherichia coli, and ddl gene-targeted species-specific primers for Enterococcus faecalis and feces Enterococcus were designed. Patients with colorectal cancer (colorectal cancer group, n=30) and healthy volunteers (normal control group, n=30) were included and whose feces were collected to extract bacterial genome DNA. SYBR GreenⅠ real-time fluorescence quantitative PCR was used to analyze the five mentioned bacterial amounts. Results Level of Bifidobacterium spp. (4.52±0.49) and Lactobacillus group (5.46±0.12) in colorectal cancer group were significantly lower than those （9.25±0.83 and 7.45±0.37） of normal control group (Plt;0.05), whereas levels of Escherichia coli (5.82±0.47), Enterococcus faecalis (10.6±0.30) and feces Enterococcus (5.74±0.16) in colorectal cancer group were significantly higher than those （4.68±0.32, 4.95±0.24, and 5.03±0.43） of normal control group (Plt;0.05). Conclusions The fecal microflora composition of patients with colorectal cancer is significantly decreased in Bifidobacterium spp. and Lactobacillus group, whereas increased in Escherichia coli, Enterococcus faecalis, and feces Enterococcus. These data underline that the occurrence and progress of colorectal cancer may be related to intestinal microflora.
ObjectiveTo evaluate diagnostic performance of crithidia luciliae immunofluorescence test (CLIFT), enzyme linked immunosorbent assay (ELISA), linear immunoassay (LIA) and chemiluminescence immunoassay (CLIA) for detection of anti-dsDNA antibodies for systemic lupus erythematosus (SLE). MethodsA total of 178 sera[SLE (n=86), other systemic rheumatic diseases (n=62), and healthy individual (n=30)], from whom received treatment from July 2012 to June 2013, were tested by 4 different assay kits. ResultsThe diagnostic performances of four methods for detecting anti-dsDNA antibodies for SLE were ELISA, CLIA, CLIFT and LIA, from higher to lower; while ELISA had the highest sensitivity (67.4%), and CLIA had the highest specificity (95.6%). The three test methods (ELISA, LIA, CLIA) had almost perfect concordance with the comparison method (CLIFT, Kappa >0.8). With cut-off values set at 95% of specificity, there was no statistical difference of sensitivity between ELISA and CLIA (58.1%, 60.5%; P>0.05). ConclusionFour assays can be used for the clinical detection of anti-dsDNA antibodies, and the results have an almost perfect concordance. Different assays show various performances depending on the methods and cut-off values used.
ObjectiveTo verify the consistency between artificial interpretation and automatic interpretation by HELIOS automatic immunofluorescence system by comparing their results on the same antinuclear antibodies (ANA) fluorescent slides, and analyze the application of automatic interpretation clinically. MethodA total of 281 ANA fluorescent slides of 281 impatients or outpatients in February 2015 were analyzed by HELIOS automatic immunofluorescence system and artificial interpretation respectively. As HELIOS could only determine the titer not the fluorescence type, only the negative or positive results qualitatively and the titer of ANA positive slides were analyzed. ResultsThere was no statistically significant difference between HELIOS automatic immunofluorescence system and artificial interpretation in negative or positive rate qualitatively (P>0.05) . The total coincidence rate was 98.9%, the positive coincidence rate was 99.5%, and the negative coincidence rate was 97.4%, and the kappa coefficient was 0.973. The difference of titer between the two groups had no statistical significance (P>0.05) . ConclusionsThe results of HELIOS automatic Immunofluorescence system and artificial interpretation are in good consistency. HELIOS automatic immunofluorescence system is suitable for clinical use as its high degree of automation, simple operation and result reliability.
This paper presents a surgical optical navigation system with non-invasive, real-time, and positioning characteristics for open surgical procedure. The design was based on the principle of near-infrared fluorescence molecular imaging. The in vivo fluorescence excitation technology, multi-channel spectral camera technology and image fusion software technology were used. Visible and near-infrared light ring LED excitation source, multi-channel band pass filters, spectral camera 2 CCD optical sensor technology and computer systems were integrated, and, as a result, a new surgical optical navigation system was successfully developed. When the near-infrared fluorescence was injected, the system could display anatomical images of the tissue surface and near-infrared fluorescent functional images of surgical field simultaneously. The system can identify the lymphatic vessels, lymph node, tumor edge which doctor cannot find out with naked eye intra-operatively. Our research will guide effectively the surgeon to remove the tumor tissue to improve significantly the success rate of surgery. The technologies have obtained a national patent, with patent No. ZI.2011 1 0292374.1.
Immuno-fluorescence technique can qualitatively determine certain nuclear translocation, of which NF-κB/p65 implicates the activation of NF-κB signal pathways. Immuno-fluorescence analysis software with independent property rights is able to quantitatively analyze dynamic location of NF-κB/p65 by computing relative fluorescence units in nuclei and cytoplasm. We verified the quantitative analysis by Western Blot. When we applied the software to analysis of nuclear translocation in lipopolysaccharide (LPS) induced (0.5 h, 1 h, 2 h, 4 h) primary human umbilical vein endothelial cells (HUVECs), we found that nuclear translocation peak showed up at 2h as with calculated Western blot verification results, indicating that the inventive immuno-fluorescence analysis software can be applied to the quantitative analysis of immuno-fluorescence.