Objective:To compare the differentiation efficiency of human induced pluripotent stem cell (hiPSC)-derived retinal pigment epithelium (RPE) cells using the Nuwacell ? RPE kit and nicotinamide (NIC)-based differentiation methods. Methods:Normal hiPSC lines and hiPSC lines from peripheral blood of two Bietti crystalline corneoretinal dystrophy (BCD) patients with CYP4V2 gene mutation at passages 16-18, were cultured for 4 days, and were induced into Nuwacell-RPE and NIC-RPE cells using the kit-based differentiation method and the NIC differentiation method, respectively.The percentage of pigmented cells or pigment foci areas formed at 3, 4, and 5 weeks after induction was observed and quantified under a light microscope.The cellular pigmentation level was measured by spectrophotometry.The expression of maturation markers in RPE cells was detected by real-time quantitative PCR and immunofluorescence staining.Cell polarity structures were examined by transmission electron microscopy.The induced RPE cells were co-cultured with porcine photoreceptor outer segments for 48 hours, and the number of rhodopsin (RHO)-labeled POS phagocytosed was quantified by immunofluorescence staining.Transepithelial electrical resistance was measured to evaluate the barrier function of the cells.According to the approval of the Ethics Committee of the First Affiliated Hospital of Army Medical University of China (No.[A]KY2023007), and peripheral blood from two BCD patients with CYP4V2 gene mutation was collected with written informed consent. Results:The Nuwacell method formed pigmented cells at 3 weeks, with (54.513±5.795)% pigmentation at 5 weeks, while the NIC method formed pigment spots at 4 weeks, with (26.037±8.489)% pigmentation at 5 weeks, with a statistically significant difference ( P<0.005).After 30 days, both methods produced hexagonal RPE structures, with pigmentation levels of (40.060±4.076) and (57.292±2.588)pg/cell, respectively, and the difference was statistically significant ( P<0.001).PCR and immunofluorescence staining showed that increased expression of maturation marker genes RPE65, BEST1, and CRALBP induced by the two methods.NIC-RPE cells phagocytosed more RHO-labeled outer segments than Nuwacell-RPE, showing a statistically significant difference ( t=2.920, P=0.043).After 4 weeks of culture in Transwell chambers, the transepithelial electrical resistance of NIC-RPE was higher than Nuwacell-RPE cells, demonstrating a statistically significant difference ( P<0.05).After continuous passage for 4 generations, Nuwacell-RPE partially lost its original cellular characteristics, including loss of pigmentation, irregular shape and loss of RPE marker expression, while NIC-RPE maintained its original morphologic features. Conclusions:The Nuwacell kit method has higher differentiation efficiency and RPE cell yield than the NIC-directed differentiation method, and cell maturity, phagocytosis, and barrier functions are inferior to those of the NIC-directed differentiation method.

Objective:To compare the differentiation efficiency of human induced pluripotent stem cell (hiPSC)-derived retinal pigment epithelium (RPE) cells using the Nuwacell ? RPE kit and nicotinamide (NIC)-based differentiation methods. Methods:Normal hiPSC lines and hiPSC lines from peripheral blood of two Bietti crystalline corneoretinal dystrophy (BCD) patients with CYP4V2 gene mutation at passages 16-18, were cultured for 4 days, and were induced into Nuwacell-RPE and NIC-RPE cells using the kit-based differentiation method and the NIC differentiation method, respectively.The percentage of pigmented cells or pigment foci areas formed at 3, 4, and 5 weeks after induction was observed and quantified under a light microscope.The cellular pigmentation level was measured by spectrophotometry.The expression of maturation markers in RPE cells was detected by real-time quantitative PCR and immunofluorescence staining.Cell polarity structures were examined by transmission electron microscopy.The induced RPE cells were co-cultured with porcine photoreceptor outer segments for 48 hours, and the number of rhodopsin (RHO)-labeled POS phagocytosed was quantified by immunofluorescence staining.Transepithelial electrical resistance was measured to evaluate the barrier function of the cells.According to the approval of the Ethics Committee of the First Affiliated Hospital of Army Medical University of China (No.[A]KY2023007), and peripheral blood from two BCD patients with CYP4V2 gene mutation was collected with written informed consent. Results:The Nuwacell method formed pigmented cells at 3 weeks, with (54.513±5.795)% pigmentation at 5 weeks, while the NIC method formed pigment spots at 4 weeks, with (26.037±8.489)% pigmentation at 5 weeks, with a statistically significant difference ( P<0.005).After 30 days, both methods produced hexagonal RPE structures, with pigmentation levels of (40.060±4.076) and (57.292±2.588)pg/cell, respectively, and the difference was statistically significant ( P<0.001).PCR and immunofluorescence staining showed that increased expression of maturation marker genes RPE65, BEST1, and CRALBP induced by the two methods.NIC-RPE cells phagocytosed more RHO-labeled outer segments than Nuwacell-RPE, showing a statistically significant difference ( t=2.920, P=0.043).After 4 weeks of culture in Transwell chambers, the transepithelial electrical resistance of NIC-RPE was higher than Nuwacell-RPE cells, demonstrating a statistically significant difference ( P<0.05).After continuous passage for 4 generations, Nuwacell-RPE partially lost its original cellular characteristics, including loss of pigmentation, irregular shape and loss of RPE marker expression, while NIC-RPE maintained its original morphologic features. Conclusions:The Nuwacell kit method has higher differentiation efficiency and RPE cell yield than the NIC-directed differentiation method, and cell maturity, phagocytosis, and barrier functions are inferior to those of the NIC-directed differentiation method.

Objective:To analyze the relationship between stress hyperglycemia (SHG) and the prognosis of acute myocardial infarction (AMI) without diabetes mellitus (DM).Methods:Using a retrospective cohort study method, 396 AMI patients without DM or impaired glucose tolerance (IGT) and admitted glycated hemoglobin A 1c (HbA 1c)≤6.0% from January 2018 to December 2020 in Xuanwu Hospital, Capital Medical University were selected. Among them, 238 patients did not have SHG at admission (group A), 85 patients had SHG at admission but their blood glucose level did not reach the diagnostic criteria for DM (group B), and 73 patients had SHG at admission and their blood glucose level reached the diagnostic criteria for DM (group C). The baseline data and the incidence of main adverse cardiovascular and cerebrovascular events (MACCE) were recorded. Multivariate Cox regression was used to analyze the independent risk factors of MACCE after discharge in AMI patients without DM. Results:The incidence of MACCE after discharge in group B and group C was significantly higher than that in group A: 29.4% (25/85) and 35.6% (26/73) vs. 18.5% (44/238), the incidence in group C was significantly higher than that in group B, and there was statistical difference ( P<0.05). Multivariate Cox regression analysis result showed that SHG on admission was an independent risk factor for MACCE after discharge in AMI patients without DM ( P<0.05), and LVEF on admission was an independent protective factor for MACCE after discharge in AMI patients without DM ( P<0.01). Conclusions:SHG on admission is the independent risk factor of MACCE in AMI patient without DM. Early detection, assessment and proper intervention measures based on clinical reality should be advocated for the AMI patients with SHG to further improve the prognosis.

BACKGROUND: Although previous studies have shown that meteorological factors such as temperature are related to the incidence of bacillary dysentery (BD), researches about the non-linear and interaction effect among meteorological variables remain limited. The objective of this study was to analyze the effects of temperature and other meteorological variables on BD in Beijing-Tianjin-Hebei region, which is a high-risk area for BD distribution. METHODS: Our study was based on the daily-scale data of BD cases and meteorological variables from 2014 to 2019, using generalized additive model (GAM) to explore the relationship between meteorological variables and BD cases and distributed lag non-linear model (DLNM) to analyze the lag and cumulative effects. The interaction effects and stratified analysis were developed by the GAM. RESULTS: A total of 147,001 cases were reported from 2014 to 2019. The relationship between temperature and BD was approximately liner above 0 °C, but the turning point of total temperature effect was 10 °C. Results of DLNM indicated that the effect of high temperature was significant on lag 5d and lag 6d, and the lag effect showed that each 5 °C rise caused a 3% [Relative risk (RR) = 1.03, 95% Confidence interval (CI): 1.02-1.05] increase in BD cases. The cumulative BD cases delayed by 7 days increased by 31% for each 5 °C rise in temperature above 10 °C (RR = 1.31, 95% CI: 1.30-1.33). The interaction effects and stratified analysis manifested that the incidence of BD was highest in hot and humid climates. CONCLUSIONS This study suggests that temperature can significantly affect the incidence of BD, and its effect can be enhanced by humidity and precipitation, which means that the hot and humid environment positively increases the incidence of BD.
Beijing/epidemiology* ; China/epidemiology* ; Dysentery, Bacillary/epidemiology* ; Humans ; Humidity ; Temperature

Objective:To explore the value of using metagenomic next-generation sequencing (mNGS) technology to detect pathogens in patients with burns and patients with acute or chronic wounds.Methods:A retrospective observational study was conducted. From March 2019 to June 2020, 11 patients with burns and patients with acute or chronic wounds (including 10 males and 1 female, aged 23 to 85 years) in the Fourth Medical Center of PLA General Hospital met the inclusion criteria and were recruited. A total of 23 specimens were collected, including 6 whole blood specimens, 1 skin tissue specimen, 1 drained pus specimen, and 15 wound secretion swab specimens. Each specimen was divided into two parts, which were subjected for pathogen detection using microbial culture method and mNGS method, respectively. The number and types of pathogens detected by the 2 methods and the relative abundance detected by the mNGS method were recorded, and the consistency of the two methods were compared. Data were statistically analyzed with paired Wilcoxon rank sum test.Results:With the microbial culture method, no pathogen was detected in 5 of the 23 specimens, while 35 pathogens were detected in the remaining 18 specimens, belonging to 9 species of bacteria and 2 species of fungi. Five specimens had one pathogen while 9 specimens had 2 pathogens and 4 specimens had 3 pathogens detected in each specimen. With the mNGS method, no pathogen was detected in one of the 23 specimens, while 75 pathogens were detected in the remaining 22 specimens, belonging to 28 species of bacteria, 3 species of fungi, and 3 species of viruses. Eight specimens had one pathogen, 5 specimens had 2 pathogens, 2 specimens had 3 pathogens, 3 specimens had 4 pathogens, 2 specimens had 6 pathogens, and 1 specimen had 7 pathogens, and 1 specimen had 20 pathogens detected in each specimen. The number of pathogens detected in each specimen by microbial culture method was 2 (1, 2) types, which was significantly less than 2 (1, 4) types by mNGS method ( Z=3.359, P<0.01). In 5 specimens, no bacteria were detected by microbial culture method but mNGS method detected bacteria in 2 specimens and virus in 2 different specimens. The mNGS method detected two or more types of bacteria in 13 specimens, the relative abundance of bacteria with the 1st relative abundance ranking ranged from 28.8% to 95.9% in each specimen. Of the 23 specimens detected by two detection methods, 7 specimens (30.4%) showed identical detection results, 5 specimens (21.7%) showed totally different detection results, and 11 specimens (47.8%) had partially consistent detection results. Conclusions:Compared with the traditional microbial culture method, the mNGS method has higher detection sensitivity and stronger capacity to detect pathogens, and can determine the relative abundance of pathogens in mixed infections. As a supplement to the culture method, the mNGS method is expected to play an important role in the diagnosis of infectious pathogens in burns and acute or chronic wounds.

From July 2018 to April 2019, 10 patients (8 males and 2 females, aged 5 to 53 years) who need skin grafting with 17 hands were admitted to the Fourth Medical Center of PLA General Hospital. The method of glove bandaging was used for wound hemostasis in skin grafting. After the initial hemostasis, sterile latex gloves were used as the inner cover of hand, and then pressure-wrapped with gauze. After 20 minutes, the gauze and gloves were removed in order from the near to the far, then thorough hemostasis was conducted again, and finally skin grafting was performed. The use of gloves as inner cover can avoid the adhesion of gauze fibers to the wound surface, so that it will not cause small blood vessels to re-bleed when the gauze is removed, therefore reducing the excessive use of electrocoagulation and secondary damage to the wound. As a result, blood loss can be reduced and hemostatic time can be shortened.

Objective: To observe the clinical effects of stage-Ⅱ Meek skin grafting on adipose tissue after tangential excision in patients with extensive deep burns, and to explore the functional mechanism. Methods: The medical records of 26 extensively burned patients who met the inclusion criteria and were admitted to the Department of Burns and Plastic Surgery of the Fourth Medical Center of PLA General Hospital from May 2015 to December 2017 were retrospectively analyzed. According to the treatment methods, 14 patients were enrolled in stage-Ⅰ skin grafting group (10 males and 4 females, aged 27 to 75 years), and 12 patients were enrolled in stage-Ⅱ skin grafting group (10 males and 2 females, aged 31 to 76 years). Patients in the 2 groups all underwent debridement of tangential excision, and their healthy adipose tissue was preserved. Meek skin grafting was performed just after tangential excision in patients in stage-Ⅰ skin grafting group. In patients in stage-Ⅱ skin grafting group, porcine acellular dermal matrix (ADM) was applied to cover the wound after tangential excision, and 3 days later, it was removed and Meek skin grafting was performed. The times of complement skin grafting and the wound basic healing time of patients in the 2 groups were observed and recorded. In the stage-Ⅱ skin grafting group, the adipose tissue of patients were taken from the wound center immediately after tangential excision and immediately after the removal of porcine ADM, for the observation of structure of the fault surface of adipose tissue through hematoxylin and eosin staining and microvessel density (MVD) through immunohistochemical staining. Data were processed with independent sample t test and Fisher′s exact probability test. Results: (1) The times of complement skin grafting of patients in stage-Ⅱ skin grafting group was (1.83±0.17) times, which was obviously less than (3.36±0.63) times in stage-Ⅰ skin grafting group (t=2.19, P<0.05). The wound basic healing time of patients in stage-Ⅱ skin grafting group was (35.1±2.3) d, which was obviously shorter than (48.8±4.9) d in stage-Ⅰ skin grafting group (t=2.27, P<0.05). (2) Immediately after tangential excision, the intercellular substance was few between the adipose cells in adipose tissue of patients in stage-Ⅱ skin grafting group. Immediately after the removal of porcine ADM, there was regenerated granulation tissue in the intercellular space of adipose cells of adipose tissue of patients in stage-Ⅱ skin grafting group. Immediately after tangential excision, the MVD of adipose tissue of patients in stage-Ⅱ skin grafting group was 20.2±1.3 under per 400-time field, which was obviously less than 32.2±1.9 under per 400-time field immediately after the removal of porcine ADM (t=-5.38, P<0.01). Conclusions Meek skin grafting on the adipose tissue in stage-Ⅱ surgery after tangential excision could reduce the times of complement skin grafting and shorten wound healing time of patients with extensive deep burns. The mechanism may be related to the improvement of the recipient condition of adipose tissue.

Objective To study the effect and mechanism of dexmedetomidine combined with sufentanil on postoperative analgesia in elderly patients with abdominal surgery. Methods Ninety-six elderly patients having undergone abdominal surgery in the First Affiliated Hospital of Wenzhou Medical University from January 2016 to June 2017 were enrolled, and they were divided into a control group and an observation group by random number table method, 48 cases in each group. General anesthesia was performed in the operation, and after surgery venous analgesic pump was used for analgesia in both groups. Analgesic method: the control group was given sufentanil 2 μg/kg and tropisetron 5 mg; the observation group was given dexmedetomidine 2 μg/kg, sufentanil 2 μg/kg and tropisetron 5 mg. The changes of pain and sedation score, conventional indexes [systolic blood pressure (SBP), diastolic blood pressure (DBP), heart rate (HR), respiratory rate (RR), pulse blood oxygen saturation (SpO2)], oxidative damage indexes [lipid peroxide (LPO), glutathione peroxidase (GSH-Px), Cu-Zn superoxide dismutase (Cu-Zn SOD)], stress response indexes [cortisol (Cor), epinephrine, norepinephrine (NE)], platelet activation indexes granule membrane protein-140 (GMP-140), thromboxane A2(TXA2) and the incidence of adverse reactions were observed in both groups. Results ① After surgery the visual simulation score (VAS) and Ramsay score in both groups were higher than those before surgery, and showed a tendency firstly increased and then decreased, and reached to peak value 2 hours after operation [VAS score:the control group was 3.24±0.98 vs. 1.95±0.93, observation group was 3.19±1.03 vs. 1.98±0.95; Ramsay score:the control group was 3.26±0.51 vs. 1.90±0.45, observation group was 3.77±0.53 vs. 1.92±0.42], began to decline 6 hours after operation, reached to valley value 48 hours after operation, and there was no significant difference in VAS scores between the two groups (2.02±0.64 vs. 1.98±0.95), Ramsay score was significantly higher in observation group than that in control group (2.59±0.41 vs. 2.10±0.21). ② Since 2 hours after the operation, the SBP, DBP, HR and RR in the observation group began to be lower than those in control group, 12 hours after surgery their values reached their valleys [SBP (mmHg, 1 mm Hg = 0.133 kPa): 113.2±13.5 vs. 122.1±10.3, DBP (mmHg): 67.5±9.9 vs. 76.4±8.6, HR (bpm): 64.5±6.9 vs. 71.4±7.5, RR (bpm): 14.8±1.1 vs. 15.8±0.8, all P < 0.05]; while SpO2did not change a great deal. ③ LPO, Cor, epinephrine, NE, GMP-140, TXA2of two groups after operation were higher than those before operation, GSH-Px and Cu-Zn SOD were lower than those before operation. However, LPO, Cor, epinephrine, NE, GMP-140, TXA2in observation group were significantly lower than those in control group [LPO (μmol/L): 6.4±0.8 vs. 9.8±1.1, Cor (ng/L): 148.2±19.1 vs. 239.3±27.8, epinephrine (μg/L): 124.2±13.9 vs. 207.1±23.5, NE (μg/L): 109.2±14.8 vs. 183.3±21.6, GMP-140 (μg/L): 27.13±3.82 vs. 39.06±4.83, TXA2(ng/L): 422.30±53.74 vs. 610.43±73.21, all P < 0.05], GSH-Px and Cu-Zn SOD in observation group were significantly higher than those in the control group [GSH-Px (U/L): 426.7±58.7 vs. 307.9±51.2, Cu-ZnSOD (μg/L): 311.3±42.5 vs. 231.6±34.1, all P < 0.05].④ The incidence of adverse reaction nausea in the observation group was significantly lower than that in the control group [4.17% (2/48) vs. 37.5% (18/48)]. Conclusion Dexmedetomidine combined with sufentanil used in elderly patients after abdominal surgery has significant analgesic effect, can effectively inhibit platelet activation, and decrease the contents of GMP-140 and TXA2.

Objective: To observe the effects of minimally invasive tangential excision in treating deep partial-thickness burn wounds on trunk and limbs in pediatric patients in the early stage post burn. Methods: Clinical data of 40 children with deep partial-thickness burn wounds on trunk and limbs, admitted to our burn ward from January 2016 to June 2017, conforming to the study criteria, were retrospectively analyzed. They were divided into conventional treatment group (CT, n=19) and minimally invasive tangential excision group (MITE, n=21) according to the different treatments. The patients in group CT were treated with eschar-reserving therapy firstly. When tangential excision was performed, the roller knife was used, and no necrotic tissue left on the wound bed was considered the proper depth of excision. Razor-thickness skin grafting was performed to cover the wound when adipose tissue exposed markedly after tangential excision. Dressing change was performed within 48 h after the operation and repeated every 2 days. Unhealed wounds were covered by razor-thickness skin grafting. The patients in group MITE were treated with tangential excision in the early stage post burn. The tangential excision was operated with electric dermatome, and the thickness was set at 0.1 mm to excise the surface of eschar until the sporadic punctate hemorrhage on wound surface was observed and some necrotic tissue was left on the wound bed. Porcine acellular dermal matrix was applied after tangential excision. The first dressing change was often performed about 1 week after the operation. Razor-thickness skin grafting was performed to cover the unhealed wounds. The length of wound healing, high fever, antibiotic usage, and hospital stay, times of later operation, and hospitalization expenses of patients in the 2 groups were recorded. The excisional eschar and wound bed tissue of patients in group MITE were harvested for pathological observation. Data were processed with t test and Fisher′s exact probability test. Results: (1) There were no statistically significant differences in length of high fever and length of hospital stay and hospitalization expenses between patients in the 2 groups (t=-1.67, -1.93, 0.31, P>0.05). The lengths of wound healing [(24.8±2.5) d] and antibiotic usage [(4.4±0.7) d] of patients in group MITE were significantly shorter than those in group CT [(33.3±2.5) and (7.0±0.7) d, t=-2.44, -2.44, P<0.05], and times of later operation of patients in group MITE [(0.29±0.14) times] were significantly less than those in group CT [(0.79±0.21) times, t=-2.03, P<0.05]. (2) The thickness of the excisional eschar of patients in group MITE was about 150 μm. The eschar has epidermis and upper dermis. Some necrotic tissue was left on the wound bed. Conclusions The treatment for pediatric deep partial-thickness burn wounds on trunk and limbs with minimally invasive tangential excision using electric dermatome in the early stage post burn can accelerate wound healing, shorten length of antibiotic usage, and reduce times of later operations.

Objective To compare the performance of 2％ (w/v) agarose gel,2％ (w/v) Metaphor agarose gel and 10％(w/v) nondenaturating polyacrylamide gel in the PCR-restriction fragment length polymorphism (RFLP) analysis of mycobacterial heat shock protein 65 (hsp65) gene.Methods This study included 8 Mycobacteria strains,including clinical isolates and standard strains of Mycobacteria tuberculosis and Mycobacterium intracellulare.Bacterialsuspension of these strains was prepared with the concentration of bacterial cells varying from 10 to 106per milliliter.PCR was performed to amplify the hsp65 gene with a pair of universal primers followed by the digestion of amplicons with two restriction endonucleases,BstE Ⅱ and Hae Ⅲ.Then,the restriction enzyme-digested fragments were subjected to electrophoresis in 2％ agarose gel,2％ Metaphor agarose gel and 10％ nondenaturating polyacrylamide gel respectively.Results As analysis of variance showed,the three gel electrophoresis methods were statistically different in sensitivity for the RFLP analysis of mycobacterial hsp65 gene (F =36.379,P ＜ 0.01).Least significance difference (LSD) procedure revealed that the 2％ agarose gel-based electrophoresis was less sensitive than the 2％ Metaphor agarose gel-and 10％ non-denaturing polyacrylamide gel-based electrophoresis (both P ＜ 0.01),and no significant differences were observed between the 2％ Metaphor agarose gel-and 10％ non-denaturing polyacrylamide gel-based electrophoresis (P ＞ 0.05).Conclusion The 2％ Metaphor agarose gel-and 10％nondenaturating polyacrylamide gel-based electrophoresis methods appear to be more sensitive than the 2％ agarose gel-based electrophoresis method for the PCR-RFLP analysis of mycobacterial hsp65 gene.