Here we reviewed the applications of laser scanning confocal microscope in biomedical field. The reformation and improvement based on this technique were also summarized. After analyzing the defects, we highlighted its potential applications in the future.

Objective To study effects of Kanglaite injection combined with gefitinib on inflammatory factors and immune function in patients with advanced lung cancer.Methods 98 patients with advanced lung cancer in First People 's Hospital of Yuhang District from December 2013 to September 2016 were selected and randomly divided into gefitinib-treated group and combination drug-treated group with 49 cases in each group.The patients in the gefitinib-treated group were treated with gefitinib, the patients in the combination drug-treated group were treated with gefitinib and Kanglaite injection.The clinical efficacy, inflammatory factors, immune function, quality of life and adverse reactions were observed and compared between the two groups. Results The efficacy and control rate in combination drug-treated group were higher than gefitinib-treated group(P<0.05).The inflammatory factors such as CRP, IL-6 and TNF-αwere significantly improved in the two groups before treatment, and the combination drug-treated group improved significantly better than gefitinib-treated group(P<0.05).The levels of cellular immune factors in the two groups were significantly improved compared with that before treatment, and the combination drug-treated group was significantly better than the gefitinib-treated group(P<0.05).The quality of life in the combination drug-treated group was significantly higher than the gefitinib-treated group(P<0.05).The incidence of adverse reactions such as nausea and vomiting, thrombocytopenia, and renal dysfunction in combination drug-treated group was lower than in the gefitinib-treated group( P<0.05).Conclusion Application of gefitinib combined with Kanglaite injection in the treatment of patients with advanced lung cancer treatment is more significant, can effectively reduce the systemic inflammation and adverse reactions, improve immune function and quality of life.

An in vitro detection method of the gastrointestinal absorption of Pilose Antler protein was established for mixed protein activity. Five bands of protein with molecular weight of 17.8-160 kD derived from the Pilose Antler were extracted and sufficiently labeled with FITC (FITC-PE). The stability and variation of FITC-PE in gastrointestinal circumstances were detected by native polyacrylamide gel electrophoresis and confocal laser scanning microscope. Results showed that the main component of FITC-PE kept invariant after being reacted with artificial gastric fluid and artificial intestinal fluid. The fluorescence signal was detected 20 min after administration in the valgus intestinal purse experiment, and three kinds of protein, with molecular weight of 45, 25, and 17.8 kD, were detected in the mixture of absorbent protein. The research laid the foundation for the further in vivo study of Pilose Antler protein. Meanwhile, it would be an in vitro screening method for the absorption, distribution and metabolism of mixed protein from traditional Chinese medicine.

Aim To establish a new,multi-parameter,real time and qualitative cell migration evaluation method.Methods Lung cancer cell line A549 was cultured on the glass bottom dish.After treated with different dosages of berberine or Rg3 for 24 hours,several scratching lines at the same dimension were made and observed by Living Cell Working Station.8 observation areas were selected randomly and imaged continuously for 24 hours.Transferred Area(TA),Transferred Distance(TD),Single Cell Transferred Speed(SCTS)and the Cell Division Number among defined area(CDN)were analyzed after getting sequence images.Results The focus stage and the incubation system were sufficient to keep cell proliferation and made it possible for long term observation.Berberine at 25 μmol·L~(-1) and 12.5 μmol·L~(-1) could inhibit the migration of A549(P<0.01).The analysis results of TA,TD,SCTS and CDN were basically coincident.Rg3 at 0.1 mmol·L~(-1) could inhibit SCTS and promote CDN in 6,12 and 24 h(P<0.01),while decrease both TA and TD in 24 hs.Conclusion The method is sensitive,rapid and simple to be applied in the research of tumor metastasis,wound healing and inflammatory response with real time,in-situ and multi-parameters.

OBJECTIVETo establish a new, real time, dynamic and direct optical detection method for mast cell degranulation caused by anaphylactoid reaction.

METHODA CD63-GFP plasmid was constructed and introduced steadily into rat basophilic leukemia (RBL-2H3) cells. The movements of CD63-GFP, which was located on both the granule membranes and the plasma membranes of RBL cells stimulated by Compound 48/80, were studied by confocal laser scanning microscope (CLSM) and total internal reflection fluorescence microscope (TIRFM) both inside and on the surface of living RBL-2H3 cells.

RESULTBefore antigen stimulation, most granules with CD63-GFP hardly moved in RBL cells. However, after antigen stimulation, the granules moved dramatically. They reached the plasma membranes in a few minutes and fused with them instantaneously. The velocity of the granule movement toward the plasma membranes on antigen stimulation was calculated to be 0.05 micron x s(-1).

CONCLUSIONAnalysis of the movement of each granule provided a new insight into the elementary process of degranulation. The method is rapid, sensitive and reliable, which could be used as a new detection method for anaphylactoid reaction in vitro.

Anaphylaxis ; diagnosis ; immunology ; metabolism ; Animals ; Antigens, CD ; genetics ; Cell Degranulation ; Cell Line, Tumor ; Cell Movement ; Mast Cells ; cytology ; immunology ; Microscopy, Confocal ; Microscopy, Fluorescence ; Platelet Membrane Glycoproteins ; genetics ; Rats ; Tetraspanin 30 ; Time Factors

This study was conducted to explore whether cholecystokinin-B/gastrin receptor (CCKBRwt) gene and its alternative splicing variant (CCKBRi4sv) are expressed in human gastric carcinomas cell line and tissues, and to find out their relationship with the development and progression of human gastric carcinoma. The mRNA expression levels of CCKBRwt and CCKBRi4sv were detected in 30 human gastric carcinomas and normal tissues adjacent to cancer, 10 gastritis specimens, 2 autopsied normal stomach specimens as well as in a gastric carcinoma cell line SGC-7901 cells. The results revealed that the transcripts of CCKBRwt and CCKBRi4sv were observed in all of the human gastric specimens tested, but only CCKBRwt was expressed in gastric cancer cell line SGC-7901 cells. The expression levels of the two receptors were not correlated with the differentiation and metastases of gastric cancers. From the results, we infer that human gastric tissues simultaneously express CCKBRwt and CCKBRi4sv, and CCKBRi4sv may have unknown physiological functions in gastric epithelial cells.
Base Sequence ; Epithelial Cells ; metabolism ; Gastric Mucosa ; metabolism ; Gastrins ; biosynthesis ; genetics ; Gastritis ; genetics ; metabolism ; Humans ; Molecular Sequence Data ; RNA, Messenger ; biosynthesis ; genetics ; Receptor, Cholecystokinin B ; biosynthesis ; genetics ; Stomach ; metabolism ; Stomach Neoplasms ; genetics ; metabolism ; Tumor Cells, Cultured

Objective:To observe the physiological effect of bi-level positive airway pressure (BiPAP) ventilation among stable chronic obstructive pulmonary disease (COPD) patients.Methods:This was a small sample size, exploratory, interventional study. A total of 10 outpatients with stable COPD were included from Department of Pulmonary and Critical Care Medicine of Zhujiang Hospital, Southern Medical University between January 2018 and December 2018. The BiPAP mode of noninvasive mechanical ventilation was adopted. The inspiratory positive airway pressure was gradually increased from 10 cmH 2O (1 cmH 2O=0.098 kPa) to 24 cmH 2O, and each time by 2 cmH 2O. The expiratory positive airway pressure remained unchanged at 4 cmH 2O. Baseline and test data were collected before and during the ventilation for comparison, including total respiratory cycle time (T tot), inspiratory time (T i), inspiratory time (T e), inspiratory tidal volume (V Ti); mouth pressure (P mo), esophageal pressure (P eso), transdiaphragmatic pressure (P di), esophageal pressure time product (PTP es), diaphragm pressure time product (PTP di), root mean square of electromyography of diaphragm (RMS), V e/RMS, inspiratory capacity (IC), the change in end-expiratory lung volume (ΔEELV) and dynamic PEEPi (PEEPi dyn). Results:All the 10 patients completed the trial. Compared to calm breathing, V Ti, V e, P mo, IC, ΔEELV score and V e/RMS increased significantly with increasing pressure levels (all P<0.05); T e only increased significantly at 20-22 cmH 2O pressure levels compared to calm breathing ( P<0.05). P di, PTP es, PTP di, RMS and RMS/RMS max decreased significantly with increasing levels (all P<0.05). PTP es and PTP di converged to 0 and no longer showed significant changes after the 18 cmH 2O pressure level. RMS and RMS/RMS max flattened out at pressure level greater than 16 cmH 2O. T i/T tot only significantly decreased at the 20 cmH 2O pressure level compared to calm breathing. PEEPi dyn showed a tendency to decrease and then increase with increasing pressure levels. Conclusion:BiPAP ventilation, at appropriate pressure levels, significantly relieves pulmonary ventilation disorders and reduces the load of respiratory muscle in patients with stable COPD.

Objective:To evaluate the role of hypoxia-inducible factor-1α (HIF-1α)/Bcl-2/E1B 19-kDa interacting protein 3 (BNIP3) signaling pathway in dexmedetomidine-induced reduction of myocardial ischemia-reperfusion (I/R)-induced brain injury in mice.Methods:Sixty clean-grade healthy male C57BL/6 mice, aged 8-10 weeks, weighting 20-30 g, were divided into 5 groups ( n=12 each) using a random number table method: sham operation group (S group), myocardial I/R group (IR group), myocardial I/R plus dexmedetomidine group (IRD group), myocardial I/R plus HIF-1α inhibitor 2ME2 group (IR-M group), and myocardial I/R plus dexmedetomidine plus HIF-1α inhibitor 2ME2 group (IRD-M group). The myocardial I/R-induced brain injury was produced by ligating the left anterior descending coronary artery for 30 min followed by 2 h of reperfusion in anesthetized mice.Dexmedetomidine 50 μg/kg was intraperitoneally injected at 5 min before ischemia in IRD group and IRD-M group.In IR-M and IRD-M groups, 2ME2 15 mg/kg was intraperitoneally injected at 5 min before ischemia.Blood samples were collected from the thoracic aorta at 2 h of reperfusion to measure the serum S-100β protein and neuron-specific enolase (NSE) concentrations.The animals were then sacrificed, brains were removed and hippocampi were obtained for determination of the apoptosis index (by TUNEL method) and expression of HIF-1α, BNIP3, Beclin-1, microtubule-associated protein 1 light chain 3 (LC3) and phosphorylated Tau protein (p-Tau) (by Western blot) and for microscopic examination of the pathological changes in hippocampal CA1 region.LC3Ⅱ/Ⅰ ratio was calculated. Results:Compared with group S, the concentrations of serum S-100β protein and NSE and apoptosis index of hippocampal neurons were significantly increased, the expression of HIF-1α, BNIP3, Beclin-1 and p-Tau was up-regulated, LC3Ⅱ/Ⅰ ratio was increased ( P<0.05), and the pathological changes in hippocampal CA1 region were aggravated in group IR.Compared with group IR, the concentrations of serum S-100β protein and NSE and apoptosis index of hippocampal neurons were significantly decreased, the expression of HIF-1α, BNIP3 and Beclin-1 was up-regulated, the expression of p-Tau was down-regulated, and LC3Ⅱ/Ⅰ ratio was increased ( P<0.05), and the pathological changes in hippocampal CA1 region were significantly attenuated in group IRD.Compared with group IRD, the concentrations of serum S-100β protein and NSE and apoptosis index of hippocampal neurons were significantly increased, the expression of p-Tau was up-regulated, the expression of HIF-1α, BNIP3 and Beclin-1 was down-regulated, LC3Ⅱ/Ⅰ ratio was decreased ( P<0.05), and the pathological changes in hippocampal CA1 region were aggravated in IR-M and IRD-M groups. Conclusions:HIF-1α/BNIP3 signaling pathway is involved in dexmedetomidine-induced reduction of myocardial I/R-induced brain injury in mice.

Objective:To explore the molecular mechanism of dexmedetomidine(Dex)protecting ischemia/reperfusion(I/R)myocardium.Methods:Wild type mice were grouped into control(Control)group,sham operation(Sham)group,WT I/R group,WT Dex group,and Dectin-1 knock out mice were grouped into KO I/R group and KO Dex group in the in vivo study(n=6).TTC stain-ing was used to determine the myocardial infarction area(%)of the above six groups of mice.HE staining and pathological analyze was used to determine the myocardial injury.Serum TNF-α,IL-6 and IL-10 levels in mice were detected by ELISA.Flow cytometry(FCM)was used to count and sort of infiltrating M2 macrophages and neutrophils in myocardium.qPCR assay was used to determine the Dectin-1 mRNA expression in the above sorted cells.Results:TTC results showed that there was no myocardial infarction in the mice of Control group and Sham group.Compared with the WT I/R group,the infarct volume was significantly lower in WT Dex group,KO I/R group and KO Dex group(P<0.05).Compared with the KO I/R group,the infarct volume was reduced in KO Dex group(P<0.05).The results of HE staining showed that the myocardial fibers of the WT I/R group of mice were disorderly arranged,with a large number of broken myocardial fibers,while the myocardial fibers of the WT Dex group,KO I/R group and KO Dex group of mice had a little breakage,the structural damage was not significant,and the myocardial arrangement was relatively neat.The degree of myocardi-al injury of mice in KO Dex group were less than that in KO I/R group mice.ELISA results showed that compared with Sham group,the serum TNF-α and IL-6 levels of the mice in WT I/R group were significantly increased,and the IL-10 level was significantly de-creased.Compared with WT I/R group,serum TNF-α and IL-6 levels of the mice in WT Dex group and KO I/R group were significant-ly decreased,and IL-10 level was significantly increased.Compared with KO I/R group,the serum TNF-α and IL-6 levels of the mice in KO Dex group were significantly decreased,and the IL-10 level was significantly increased(P<0.05).FCM cell counting results showed that compared with Sham group,a large number of M2 macrophages and neutrophils were infiltrated in the myocardium of WT I/R group of mice(P<0.05).Compared with WT I/R group,the M2 macrophages and neutrophils infiltrated in the myocardium were significantly decreased in WT Dex group,KO I/R group and KO Dex group of mice(P<0.05).While there was no significant differ-ence between the KO I/R group and the KO Dex group mice(P>0.05).qPCR results showed that compared with Sham group,the ex-pression level of Dectin-1 mRNA in the myocardial infiltrated M2 macrophages and neutrophils were significantly up-regulated in WT I/R group of mice(P<0.05).While compared with WT I/R group,the expression level of Dectin-1 mRNA in Dex group of mice was sig-nificantly lower(P<0.05).Mice in KO I/R group and KO Dex group did not express Dectin-1.Conclusion:The protective mecha-nisms of Dex preconditioning on I/R injured myocardium involves reducing the infiltrating number of M2 macrophages and neutrophils in myocardium after I/R injury,which may be achieved by inhibiting the expression of Dectin-1.

【Objective】 To compare the clinical efficacy and sagittal parameters of oblique lateral interbody fusion （OLIF） combined with posterior percutaneous internal fixation and percutaneous transforaminal endoscope-assisted posterior lumbar interbody fusion （PT-Endo-TLIF） in treating degenerative lumbar spondylolisthesis. 【Methods】 A retrospective analysis was made on 43 patients with Meyerding Ⅰ and Ⅱ° degenerative lumbar spondylolisthesis treated in our hospital from September 2017 to January 2020. Among them 23 cases were treated by OLIF， and the other 20 cases were treated by PT-Endo-TLIF. We observed and recorded the operation time， average length of hospital stay， and intraoperative blood loss， and postoperative complications of the patients. The patients were followed up 3 day， 6 and 12 months after the operation. The lumbar sagittal parameters of the two groups were compared by X-ray， CT and MRI examinations. The patients’ lower back pain was recorded for visual analogue scale （VAS）， and Oswestry disability index （ODI） was used to evaluate the clinical efficacy. 【Results】 Both groups of patients successfully completed the operation and follow-up， with the average follow-up time of 12 months. The average amount of intraoperative blood loss and operation time were significantly lower in OLIF group than in PT-Endo-TLIF group （P<0.05）. Intervertebral height increased significantly in the two groups after operation compared with pre-operation （P<0.05）. Compared with pre-operation， lumbar lordosis angle， lower lumbar lordosis angle and lumbar lordosis distribution index increased in both groups （P<0.05）， with no significant difference between them （P>0.05）. The inclination angle of L4 vertebral body and the distance between L1 vertical line and S1 in both groups were decreased compared with those before surgery （P<0.05）， but there was no significant difference between the two groups （P>0.05）. The inclination angle of L5 vertebral body in the two groups was increased compared with that before surgery （P<0.05）， but there was no statistical significance between both groups （P>0.05）. 【Conclusion】 OLIF surgical technique has the comparative advantages of definite curative effect， less trauma， fewer surgical complications， shorter operation time， less bleeding， and good recovery of the height of intervertebral space， which is suitable for its application among clinicians.