PurposeTo optimize the imaging parameters of clinical MRI scanner in rat pancreas imaging to improve the image quality and to provide better MRI image quality and more economical research method for imaging study of rat pancreas. Materials and Methods Twenty-four healthy male Wistar rats were randomly divided into the conventional sequence (CS) group, the adjustment sequence (AS) group and the optimization sequence (OS) group, with 8 rats in each group. The rats in the CS group were scanned with conventional parameters using a clinical MRI scanner. The principle of parameter adjustment was: parameters associated with T1WI or T2WI imaging quality (TR, TE, slice thickness, NEX, FOV and matrix) was set with four changes, and only one of the six parameters was changed in each scan, image quality was evaluated by two senior radiologists, the parameter corresponded the best image quality evaluated consistently by two radiologists were selected as the optimal imaging parameter, all the optimized parameters were set up step by step in this way which formed the imaging parameters in OS group. The pancreatic signal intensity and signal to noise ratio was compared between CS group and OS group after imaging.Results The optimized sequence parameters in clinical MRI scanner were listed below: T1WI sequence (M3D/FSPGR/15): TR 6 ms, TE 2.5 ms, slice thickness 2.0 mm, NEX 8, FOV 7 cm×7 cm, Matrix 120×120; T2WI sequence (FSE-XL/90): TR 4000 ms, TE 71 ms, slice thickness 2.0 mm, NEX 1, FOV 8 cm×8 cm, Matrix 192×160. The pancreatic SI in T1WI and T2WI sequence of the OS group were significantly higher than those in the CS group (t=5.16 and 3.80,P<0.01), while the pancreatic SNR in T1WI and T2WI sequence of the OS group were significantly higher than those in the CS group (t=5.65 and 3.26,P<0.01).Conclusion The optimized parameters can improve the imaging quality of rat pancreas MRI significantly, thus provide a reference for the related experimental study.

Objective:To observe the changes in the structure and function of the retina in diabetic patients, and preliminarily explore the changes in the characteristics of neuropathy and microvascular damage in different degrees of diabetic retinopathy (DR).Methods:A prospective controlled study. From May to December 2020, 63 eyes of 63 patients with type 2 diabetes who were recruited from the Department of Ophthalmology of Shandong Provincial Hospital and 40 healthy volunteers with age and sex matching in the same period (control group) were included in the study. All subjects underwent optical coherence tomography angiography (OCTA) and portable non-mydriatic visual electrophysiological diagnosis system RETeval. OCTA was used to measure the thickness of the retinal nerve fiber layer (pRNFL) around the optic disc, the blood flow density of theradial peripapillary capillary (RPC) around the optic disc, and the thickness of the macular ganglion cell complex (GCC). The "DR evaluation plan" mode of the RETeval device was used to perform flash electroretinogram examination, and the "DR evaluation score" measured by the system was recorded. According to the DR grading standard established in the early treatment of DR research, DR was classified. Diabetic patients were divided into non-DR (non-DR) group, mild to moderate non-proliferative DR (mNPDR) group, and severe non-proliferative DR (sNPDR) group, Proliferative DR (PDR) group, with 12, 16, 18, and 17 eyes respectively. The comparison of pRNFL thickness, GCC thickness, RPC blood flow density and "DR assessment score" between groups was performed by one-way analysis of variance; the correlation between pRNFL thickness and RPC blood flow density was analyzed by Pearson correlation analysis.Results:Compared with the control group, the overall, upper and lower thickness of the macular GCC of the affected eyes in different degrees of DR groups were significantly thinner, and the difference was statistically significant ( F=13.560, 15.840, 5.480; P<0.05). Compared with the control group, the overall pRNFL ( F=6.120), upper part ( F=6.310), lower part ( F=5.330), upper nose ( F=7.350), lower nose ( F=2.690), the upper nasal side ( F=4.780), the upper temporal side ( F=3.710), and the lower temporal side ( F=3.750) became thinner, the difference was statistically significant ( P<0.05). Correlation analysis results showed that the whole optic disc, upper part, lower part, upper nose, upper nasal side, lower nasal side, and lower temporal RPC blood flow density were positively correlated with pRNFL thickness ( r=0.260, 0.256, 0.275, 0.489, 0.444, 0.542, 0.261; P<0.01). The "DR evaluation scores" of the eyes in the control group, non-DR group, mNPDR group, sNPDR group, and PDR group were 12.71±5.62, 22.18±3.77, 24.68±2.41, 24.98±2.78, 29.17±7.98 points; the DR lesions were more severe, the evaluation score were higher, and the difference was statistically significant ( F=1.535, P<0.01). Conclusion:Compared with the control group, the macular GCC, pRNFL thickness and RPC blood flow density of diabetic patients are significantly reduced; the "DR evaluation score" is increased, and it is related to the severity of DR.

OBJECTIVETo investigate the suitable transfection condition of human epidermal cells (hECs) with human epidermal growth factor (EGF) gene by adenovirus vector (Ad-hEGF) and its effects on the biological characteristics of hECs.

METHODShECs were isolated from deprecated human fresh prepuce tissue of circumcision by enzyme digestion method and then sub-cultured. hECs of the third passage were used in the following experiments. (1) Cells were divided into non-transfection group and 5, 20, 50, 100, 150, and 200 fold transfection groups according to the random number table (the same grouping method below), with 3 wells in each group. Cells in non-transfection group were not transfected with Ad-hEGF gene, while cells in the latter six groups were transfected with Ad-hEGF gene in multiplicities of infection (MOI) of 5, 20, 50, 100, 150, and 200 respectively. The morphology of the cells was observed with inverted phase contrast microscope, and expression of green fluorescent protein of the cells was observed with inverted fluorescence microscope at transfection hour (TH) 24, 48, and 72. (2) Another three batches of cells were collected, grouped, and treated as above, respectively. Then the transfection rate of Ad-hEGF gene was detected by flow cytometer (n=3), the mass concentration of EGF in culture supernatant of cells was detected by enzyme-linked immunosorbent assay (n=6), and the proliferation activity of cells was detected by cell counting kit 8 (CCK8) and microplate reader (n=6) at TH 24, 48, and 72, respectively. (3) Cells were collected and divided into non-transfection group and transfection group, with 6 wells in each group. Cells in non-transfection group were cultured with culture supernatant of cells without transfection, while cells in transfection group were cultured with culture supernatant of cells which were transfected with Ad-hEGF gene in the optimum MOI (50). CCK8 and microplate reader were used to measure the biological activity of EGF secreted by cells on culture day 1, 3, and 5. (4) Cells were collected and divided into non-transfection group and transfection group, with 12 wells in each group. Cells in non-transfection group were not transfected with Ad-hEGF gene, while cells in transfection group were transfected with Ad-hEGF gene in the optimum MOI (50). The expression levels of cytokeratin 14 (CK14) and CK19 of cells were measured by immunofluorescence staining at TH 24. (5) Cells were collected, grouped, and treated as in (4), with 6 wells in each group. At post scratch hour (PSH) 0 (immediately after scratch), 12, 24, and 48, the migration distance of cells was observed and measured with inverted phase contrast microscope. Data were processed with analysis of variance of factorial design, analysis of variance for repeated measurement, and LSD test.

RESULTS(1) At TH 24 and 48, morphology of cells in each transfection group and non-transfection group were similar. Compared with that in non-transfection group, the cell debris increased significantly in 200 fold transfection group at TH 72. At TH 24, 48, and 72, the expression of green fluorescent protein was not seen in cells of non-transfection group, whereas it increased in cells of transfection group over transfection time. (2) The transfection rate of Ad-hEGF gene of cells in each transfection group increased gradually over transfection time. At TH 72, the transfection rates of Ad-hEGF gene of cells in 50-200 fold transfection groups were all above 90%, while the transfection rates of Ad-hEGF gene of cells in non-transfection group, 5, and 20 fold transfection groups were (0.51±0.20)%, (62.44±6.23)%, and (75.00±5.43)% respectively, which were obviously lower than the rate in 50 fold transfection group [(93.12±2.55)%, with P values below 0.01]. The mass concentration of EGF in culture supernatant of cells in each transfection group increased gradually over transfection time. At TH 72, the mass concentration of EGF in culture supernatant of cells in 50 fold transfection group was obviously higher than that in each of the other groups (with P values below 0.01). The proliferation activity of cells in each group at TH 24 and 48 was similar (with P values above 0.05). At TH 72, the proliferation activity of cells in 200 fold transfection group was obviously lower than that in other groups (with P values below 0.05). (3) On culture day 1, the biological activity of EGF secreted by cells in two groups was similar (P>0.05). On culture day 3 and 5, the biological activity of EGF secreted by cells in transfection group were obviously higher than that in non-transfection group (with P values below 0.01). (4) At TH 24, the expression levels of CK14 and CK19 of cells in transfection group were higher than those in non-transfection group. (5) The width of scratch in two groups was nearly the same at PSH 0. At PSH 12-48, the migration distance of cells in transfection group was obviously longer than that in non-transfection group (with P values below 0.01).

CONCLUSIONSThe suitable range of MOI of hECs transfected with Ad-hEGF gene is 50-150, and 50 is the optimum. hECs transfected with Ad-hEGF gene with MOI 50 can effectively express the EGF gene and keep its good abilities of proliferation, differentiation, and migration, as well.

Adenoviridae ; Cell Differentiation ; Cell Proliferation ; Cells, Cultured ; EGF Family of Proteins ; genetics ; metabolism ; Epidermis ; cytology ; Genetic Vectors ; Humans ; Keratins ; metabolism ; Male ; Transfection

Objective To evaluate the optimum compatibility of nalbufine combined with ropivacaine for patient-controlled epidural analgesia (PCEA) after cesarean section.Methods A total of 100 parturients who were at full term with a singleton fetus,aged 24-35 yr,with body mass index of 29-33 kg/m2,of American society of Anesthesiologists physical status Ⅱ,scheduled for elective cesarean section under combined spinal-epidural anesthesia,were divided into 4 groups (n =25 each) using a random number table method:sufentanil 0.5 μg/ml plus 0.15％ ropivacaine group (SR group),nalbufine at final concentration of 0.2 mg/ml plus 0.15％ ropivacaine group (N1 R group),nalbufine at final concentration of 0.4 mg/ml plus 0.15％ ropivacaine group (N2R group) and nalbufine at final concentration of 0.4 mg/ml plus 0.1％ ropivacaine group (N3Rgroup).PCEA solution was prepared correspondingly after surgery,and all the drugs were diluted to 100 ml in normal saline in each group.The PCA pump was set up to deliver a 0.5 ml bolus dose with a 15-min lockout interval and background infusion at 2 ml/h.Visual analog scale scores of incisional pain and anduterine contraction pain were maintained＜4.Ramsay sedation scores were recorded at 8,12,24 and 48 h after surgery.The total pressing times of PCEA were recorded within 48 h after surgery.The development of adverse reactions such as nausea,vomiting,skin itching,numbness of lower extremity,urinary retention and respiratory depression was recorded in the analgesia period.Venous blood samples were collected before surgery and at 24 and 48 h after surgery for determination of plasma prolactin concentrations,and the time of colostrum was recorded.Neonatal nerve and adaptive capacity was assessed and scored.Results Compared with group SR,the total pressing times of PCEA were significantly reduced in N2R and N3R groups (P＜0.05),and no significant change was found in the total pressing times of PCEA in group N1R (P＞0.05).Compared with group N1R,the total pressing times of PCEA were significantly reduced in N2R and N3 R groups (P＜0.05).There was no significant difference in the total pressing times of PCEA between group N2R and group N3R (P＞0.05).The Ramsay sedation score was 2 in four groups.There was no significant difference in numbness of lower extremity,plasma prolactin concentrations or neonatal nerve and adaptive capacity scores among four groups (P＞0.05).Conclusion Nalbufine at final concentraction of 0.4 mg/ml mixed with 0.1％ ropivacaine is the optimum compatibility when used for PCEA after cesarean section.