For construction of anti-CD3 human/murine chimeric antibody genes, a selective first-strand cDNA synthesis from mRNA or RNA of murine McAb HIT3a was performed using murine Ig constant region primers, and then cDNA of heavy and light chain variable domains of murine immunoglobulin were amplified by PCR using a set of degenerated oligonucleotide primers. Using these cDNA fragments as probes, the L and H chain V region exons encoding the murine McAb anti-CD3 were isolated from the gene library of HIT3a DNA and inserted into mammalian expression vectors containing the human ? and yl constant region exons for construction of human/murine chimeric antibody genes.

Objective:To screen the time points of high survival rate and efferocytosis dysfunction of rat alveolar macrophages stimulated by cigarette smoke extract (CSE), establish an in vitro model of alveolar macrophage efferocytosis function, and study chronic respiratory diseases with chronic inflammatory reaction as the main pathological changes. Methods:① Time point screening experiment: rat alveolar macrophages (NR8383 cells) were cultured in vitro, and the cells in logarithmic growth phase were divided into blank control group (100 μL complete medium) and 5% CSE group (90 μL complete medium + 10 μL 100% CSE). Alma blue method was used to detect the effect of 5% CSE on the activity of NR8383 cells at 6, 12, 24 and 48 hours. ② Apoptosis induction experiment: rat type Ⅱ alveolar epithelial cells (RLE-6TN cells) were cultured in vitro as phagocytic target cells of NR8383 cells, and the cells in logarithmic growth phase were divided into blank control group and 10, 30 and 60 minutes groups after ultraviolet exposure (apoptosis was induced by 30 000 μJ/cm 2 ultraviolet irradiation for 15 minutes). Flow cytometry was used to detect the apoptosis rate of RLE-6TN cells cultured for 10, 30 and 60 minutes after ultraviolet exposure. ③ Cell efferocytosis experiment: NR8383 cells in logarithmic phase were divided into blank control group and 5% CSE group. Two hours before NR8383 cells were stimulated by CSE for 6, 12 and 24 hours, RLE-6TN cells were exposed to ultraviolet to induce apoptosis, and the RLE-6TN cell suspension was added to NR8383 cells (the ratio of RLE-6TN cells to NR8383 cells was 5∶1). Flow cytometry was used to detect the efferocytosis rate of NR8383 cells to RLE-6TN cells at different time points treated with 5% CSE. Results:① Compared with the blank control group, the activity of NR8383 cells significantly decreased after treatment with 5% CSE for 48 hours [cell reduction rate: (68.5±4.1)% vs. (73.6±2.3)%, P < 0.05]. However, there were no significant differences when the activities of NR8383 cells treated with 5% CSE for 6, 12 and 24 hours were compared with the blank control group, so these three time points were selected for the subsequent establishment of alveolar macrophage cell efferocytosis dysfunction in vitro model experiment. ② Compared with the blank control group, the apoptosis rate of RLE-6TN cells significantly increased at 10, 30 and 60 minutes after ultraviolet exposure [(66.87±8.63)%, (85.51±2.39)%, (96.13±2.74)% vs. (9.13±3.17)%, all P < 0.01] in a time-dependent manner. Considering that it taked about 50 minutes for RLE-6TN cells to be labeled with PKH26 membrane labeling probe, 10 minutes after ultraviolet exposure was selected to label RLE-6TN cells. ③ Compared with the blank control group, the efferocytosis function of NR8383 cells was significantly decreased after treatment with 5% CSE for 12 hours [cell efferocytosis rate: (33.64±1.30)% vs. (44.02±2.71)%, P < 0.01], but there was no significant effect on the efferocytosis function of NR8383 cells at 6 hours and 24 hours. Conclusions:CSE can induce alveolar macrophage cell efferocytosis dysfunction. Based on the test results of the effect of 5% CSE on NR8383 cell activity and cell efferocytosis function, 12 hours with high survival rate and weak efferocytosis effect of NR8383 cells can be selected as the in vitro model condition of alveolar macrophage cell efferocytosis dysfunction.

Objective:To investigate the effect of the 3t sensorial saturation in the application of relieving pain and comfort due to femoral vein blood sampling in preterm infants, so as to provide the reference for the selection of clinical nursing plans.Methods:This was a quasi experimental study. A total of 110 preterm infants admitted to the neonatal unit of Shanxi Children′s Hospital from August 2021 to March 2022 were selected and divided into the control group and intervention group with 52 cases respectively by the random number table method. The control group implemented conventional care, and the intervention group implemented the 3t sensorial saturation method including taste, touch and talk on the basis of the control group. The pain and comfort of two groups were evaluated by the Premature Infant Pain Profile (PIPP) and COMFORTneo Scale at 3 min before, during, and 3 min after blood, and the heart rate and SpO 2 of the two groups were compared. Results:Finally, 52 premature infants were included in both groups. The PIPP score, the total score of the COMFORTneo Scale, the heart rate and SpO 2 were 2.00 (1.00, 3.00), 6.50 (6.00, 7.75), 4.00 (3.00, 5.00), 7.00 (6.00, 8.00), 17.00 (15.00, 19.00), 9.50 (9.00, 10.00) points, (137.29 ± 8.58), (148.31 ± 8.89), (143.06 ± 7.61) times/min, 0.980 (0.970, 0.990), 0.960 (0.950, 0.970), 0.980 (0.970, 0.990) in the intervention group, 2.00 (1.25, 3.00), 12.00 (11.00, 13.00), 7.00 (6.00, 8.00), 7.00 (6.00, 9.00), 25.00 (23.00, 27.00), 20.00 (19.00, 22.00) points, (141.54 ± 10.57), (179.71 ± 14.62), (162.00 ± 14.32) times/min, 0.980 (0.960, 0.990), 0.940 (0.920, 0.958), 0.960 (0.940, 0.978). The results of generalized estimating equation analysis showed that the PIPP score, total COMFORTneo Scale score and SpO 2 via different time points, subgroups, and subgroups with time points were statistically significant (Wald χ2 values were 16.72-2 489.71, all P<0.05). The results of two-factor repeated measures ANOVA showed that the interaction effects of heart rate via different time points, subgroups, and subgroups with time points were statistically significant ( F=253.08, 105.02, 77.17, all P<0.05). Conclusions:The 3t sensorial saturation method can effectively reduce pain during femoral vein blood sampling in preterm infants, can improve the comfort level of preterm infants, is conducive to the stabilization of vital signs in preterm infants, and is suitable for promotion and application in clinical care.