Objective To explore the optimal experiment conditions of CCK-8 and MTS for cell proliferation assays in human amniotic epithelial cells and to evaluate the cytotoxicity of these reagents. Methods Human amniotic epithelial cells (hAECs) in logarithm growth stages were prepared in different cell concentrations with DMEM/F12 and 10% FBS. The sensitivity and optimal wavelengths was determined based on the optical density (OD) measured at 450 nm and 492 nm. The optimal time was determined under the conditions of the same cell concentration and defined OD values. HAECs were treated with DMSO, CCK-8 and MTS for 1 h, 2 h, 3 h, and 4 h, respectively. 24 h later, cytotoxicity of the CCK-8 and MTS was evaluated by determination of cell proliferation and Trypan Blue staining. Results The optimal detection wavelength was 450 nm for CCK-8, and 492 nm for MTS. The sensitivity of CCK-8 was slightly lower then that of MTS. The optimal time for incubation hAECs with CCK-8 was 4 h within 1~4 h. The inhibitory on cell proliferation and cytotoxicity of CCK-8 were weaker then those of MTS. Conclusion CCK-8 is a convenient reagent with low cytotoxicity for detection of the proliferation of hAECs.

Objective:To investigate the long-term outcomes of laparoscopic ventral rectopexy (LVR) for obstructive defecation with overt pelvic structural abnormalities.Methods:The retrospective cohort study was conducted. The clinical data of 31 obstructive defecation patients with overt pelvic structural abnormalities who were admitted to the Renji Hospital of Shanghai Jiaotong University School of Medicine from June 2014 to August 2020 were collected. There were 6 males and 25 females, aged 59(range, 32?81)years. All 31 patients underwent LVR through transabdominal approach. Observation indicators: (1) the Cleveland clinic constipation score (CCCS); (2) severity of obstructive defecation; (3) patients assessment of constipation quality of life (PAC-QoL). Follow-up was conducted using telephone interview and outpatient examination up to October 2021. One professional researcher assessed the constipation symptoms and quality of life of patients through outpatient interview or mobile software platform of Questionnaire Star. Measurement data with skewed distribution were represented as M(range), and comparison before and after operation was conducted using the Wilcoxon sign rank test. Results:(1) The CCCS. All 31 patients underwent LVR for the first time and were followed up for 61.8(range, 11.0?87.0)months. The constipation symptoms of the 22 patients were improved. The CCCS of the 31 patients before surgery and at the last follow-up time were 15.8(range, 8.0?26.0) and 10.7(range, 2.0?20.0), respectively, showing a significant difference ( Z=?3.98, P<0.05). (2) Severity of obstructive defecation. The severity scores of frequency of bowel movements, difficult of bowel movements, sensation of incomplete defecation, abdominal distension or pain, time of each bowel movements, daily unsuccessful times of defecation, artificial assisted defecation for the 31 patients were 2.9(range, 1.0?4.0), 3.0(range, 1.0?4.0), 1.9(range, 0?3.0), 0.5(range, 0?3.0), 2.6(range, 2.0?4.0), 2.0(range, 0?4.0), 0.9 (range, 0?2.0) before surgery, versus 1.7(range, 0?4.0), 1.6(range, 0?4.0), 1.2(range, 0?4.0), 0.3(range, 0?3.0), 1.7(range, 0?3.0), 1.4(range, 0?3.0), 0.7(range, 0?2.0) after surgery, respectively. There were significant differences in the frequency of bowel movements, difficult of bowel movements, sensation of in-complete defecation, abdominal distension or pain, time of each bowel movements, daily unsuccessful times of defecation for the 31 patients before and after surgery ( Z=?3.38, ?3.80, ?2.54, ?2.31, ?3.64, ?2.75, P<0.05) and there was no significant difference in the artificial assisted defecation for the 31 patients before and after surgery ( Z=?1.31, P>0.05). (3) PAC-QoL. The score of physical discomfort, satisfaction, worries and concerns, psychological discomfort for the 31 patients were 2.3(range, 1.0?4.0), 3.2(range, 1.0?4.8), 2.2(range, 0.6?4.0), 1.8(range, 0.4?3.9) before surgery, versus 1.6(range, 0?4.0), 2.3(range, 0?4.0), 1.7(range, 0?4.0), 1.3(range, 0?4.0)after surgery, respectively, showing significant differences before and after surgery ( Z=?3.49, ?2.17, ?2.50, ?3.05, P<0.05). Conclusions:The long-term outcomes of LVR for obstructive defecation with overt pelvic structural abnorma-lities are satisfactory. Symptoms as frequency of bowel movements, difficult of bowel movements, sensation of incomplete defecation, abdominal distension or pain, time of each bowel movements and daily unsuccessful times of defecation will be significantly improved after LVR and the constipation quality of life of patients will be improved.

OBJECTIVEInflammatory reaction and injury in immature lungs are associated with activation of nuclear factor-kappa B (NF-kappaB) to trigger proinflammatory cytokine release, but the mechanism thereof is not fully understood. The present study was conducted to understand possible relationship between expression of NF-kappaB and its inhibitor and severity and outcome of neonates with hyaline membrane disease (HMD).

METHODSSerial samples of bronchoalveolar lavage fluid (BALF) were obtained during mechanical ventilation from 31 preterm infants with HMD. These infants were divided into two groups: survivors group [n = 22, birth weight (1500 +/- 320) g and gestational age (31.2 +/- 1.8) weeks] and nonsurvivors group [birth weight (1340 +/- 280) g, gestational age (30.8 +/- 2.1) weeks]. Nineteen preterm infants [birth weight (1470 +/- 280) g, gestational age (30.6 +/- 1.9) weeks] without respiratory disorders were enrolled as control subjects. Alveolar macrophages (AM) were isolated by differential adherence. AM was cultured and treated with lipopolysaccharide (LPS) for 1 hr. Then, nuclear extracts of AM were analyzed by electrophoretic mobility shift assay (EMSA) for NF-kappaB expression. NF-kappaB inhibitor (IkappaB-alpha protein) in cytoplasmic extracts was detected by using Western blotting and IL-1beta and IL-8 in BALF by enzyme-linked immunosorbent assay (ELISA).

RESULTSNF-kappaB complexes were observed by EMSA, they were characterized by competition with cold oligonucleotide and p65-specific antibodies. The addition of an excess of cold oligonucleotide, corresponding to the NF-kappaB binding site, turned off the signal of the band, showing that the band was specific. An excess of an irrelevant oligonucleotide (corresponding to the SP-1) did not show any effect. The addition of an anti-p65 antibody caused the supershift of the two upper bands. After EMSA, the NF-kappaB complexes were quantified by using a ImageQuant software. NF-kappaB expression in AM at 24 hrs was higher in all the patients with HMD as compared with control subjects (survives/control, 34.1 vs 11.4 RDU, P < 0.01; nonsurvivors/control, 55.2 vs 11.4 RDU, P < 0.01). The NF-kappaB expression in AM at 72 hrs was higher than that in control subjects but not for nonsurvivors (survivors/control, 47.8 vs 25.6 RDU, P < 0.01; nonsurvivors/control, 21.8 vs 25.6, P > 0.05). The NF-kappaB expression in AM from nonsurvivors was depressed at 72 hrs as compared to 24 hrs (21.8 vs 55.2, P < 0.01), whereas the NF-kappaB expression in AM from survivors was still higher at 72 hrs than that at 24 hrs (47.8 vs 34.1, t = 4.43, P < 0.01).

CONCLUSIONAltered NF-kappaB activation in AM of BALF of neonates with HMD was observed, and it may be mediated by decreased IkappaB synthesis, increased IkappaB degradation, or both. In HMD nonsurvivors NF-kappaB translocation was hampered upon LPS activation.

Birth Weight ; Blotting, Western ; Bronchoalveolar Lavage Fluid ; cytology ; Cell Culture Techniques ; Cell Nucleus ; drug effects ; metabolism ; Cytoplasm ; drug effects ; metabolism ; Electrophoretic Mobility Shift Assay ; Enzyme-Linked Immunosorbent Assay ; Female ; Gestational Age ; Humans ; Hyaline Membrane Disease ; immunology ; therapy ; I-kappa B Proteins ; immunology ; Infant, Newborn ; Infant, Premature ; immunology ; Interleukin-1beta ; immunology ; Interleukin-8 ; immunology ; Lipopolysaccharides ; pharmacology ; Macrophages, Alveolar ; drug effects ; immunology ; Male ; NF-KappaB Inhibitor alpha ; NF-kappa B ; immunology ; Respiration, Artificial ; Severity of Illness Index ; Time Factors

OBJECTIVETo explore the molecular mechanism and prevention of retinoic acid syndrome (RAS).

METHODSSDF-1 alpha mRNA from healthy adult lung tissue was measured by RT-PCR, CXCR4 protein expression on the cell membrane of APL cells induced by ATRA (APL-ATRA) was tested by FCM, and the rotary cell culture system (RCCS) was used to build a modal for in vitro stimulation of APL-ATRA infiltrating human lung tissue. The ability of APL-ATRA in adhesion, migration and infiltration was observed by interference from DEX, Ara-C and DNR.

RESULTSThe APL-ATRA cells could evidently infiltrate into normal lung tissue. Mean fluorescence intensity (MFI) of CXCR4 on the cell membrane of APL-ATRA cells was 30.6 +/- 1.8, which was much higher than that on unspecialized APL cells (9.8 +/- 4.2). SDF-1 alpha mRNA expression was detected positive in all 6 lung tissue. Contrary to the control groups, DEX could dramatically restrain the ability of APL-ATRA cells in adhesion and migration [(27.2 +/- 2.6)% vs. (46.0 +/- 3.0)%, (28.1 +/- 4.0)% vs. (48.2 +/- 3.0)%], while Ara-C and DNR could distinctly depress the ability in adhesion, migration and infiltration [(28.1 +/- 3.0)%, (30.2 +/- 3.2)% vs. (46.0 +/- 3.0)%; (29.0 +/- 4.0)%, (23.0 +/- 5.2)% vs. (48.2 +/- 3.0)%; (16.8 +/- 7.6)%, (17.1 +/- 6.0)% vs. (43.6 +/- 5.0)%].

CONCLUSIONIn vitro APL-ATRA cells can infiltrate into the human lung tissue. High expression of CXCR4 on APL-ATRA and SDF-1 alpha in the lung tissue may be one of the molecular mechanisms of the lung infiltration and RAS. DEX, Ara-C and DNR can dramatically restrain the ability of APL-ATRA cells in adhesion, migration and infiltration.

Adolescent ; Adult ; Cell Adhesion ; Cell Culture Techniques ; Cell Movement ; Chemokine CXCL12 ; genetics ; metabolism ; Child ; Female ; Humans ; Leukemia, Promyelocytic, Acute ; metabolism ; pathology ; Male ; Middle Aged ; Neoplasm Invasiveness ; Receptors, CXCR4 ; genetics ; metabolism ; Tretinoin ; adverse effects ; Tumor Cells, Cultured

OBJECTIVETo evaluate the effect of electromagnetic pulse (EMP) irradiation on mice reproduction.

METHODSFemale/male Kunming mice, 6 - 8 weeks old, prior to mating, or female after pregnancy were treated with whole body irradiation by 6 x 10(4) V/m electromagnetic pulse (EMP) for five times. The pregnant mice were killed on the 18th days, and teratological markers were analysed.

RESULTSEMP irradiation caused no significant changes in most of female organ weight and organ/body weight ratio. But it caused significant shortening in tail length of live foetus in the female mice before conception (prior to mating) or after pregnancy (P < 0.05), and obvious decrease in male offspring ratio (0.85 +/- 0.09 vs 1.09 +/- 0.17, P < 0.05). The male offspring ratio also significantly decreased (0.76 +/- 0.18 vs 1.09 +/- 0.17, P < 0.01) after male mice irradiated by EMP. The tail length of live foetus was shortened and male offspring sex ratio was increased after both male and female mice were irradiated by EMP. EMP irradiation also caused a significantly higher fetal death rate than normal control (P < 0.05). The embryo absorption rate was increased after irradiation except that was decreased in male mice.

CONCLUSIONEMP irradiation has effect on pregnancy and offspring development in both male and female mice before mating and in female mice after pregnancy.

Animals ; Female ; Fetus ; radiation effects ; Male ; Mice ; Pregnancy ; Radiation ; Reproduction ; radiation effects

Bronchoalveolar Lavage ; Extracorporeal Membrane Oxygenation ; Humans ; Male ; Middle Aged ; Pulmonary Alveolar Proteinosis ; therapy

OBJECTIVETo survey the prevalence of HIV-1 drug resistance after five years antiretroviral treatment in Henan province.

METHODSThrough the cross-sectional study, serum specimens of 69 HIV infected individuals that were 2 to 25 years old who were newly diagnosed according the WHO standard from November 2007 to August 2008 and did not receive antiretroviral treatment (ART) were collected. HIV-1 pol genetic mutations associated with drug resistance were identified with RT-PCR and interpreted.

RESULTSOut of 69 samples, 50 samples were successfully amplified and sequenced. Seven drug resistant mutation in reverse transcriptase region were detected and three mutations in protein region. In one specimen, a mutation (K103N) in reverse transcriptase was identified which caused high level resistance to NNRTIs, but no proteinase inhibitor mutation was found in protein region. According to the sampling and threshold surveillance criteria, the prevalence of drug resistant HIV-1 in Henan was less than 5%.

CONCLUSIONThe prevalence of drug resistant HIV-1 was still at low level in Henan. However, the proportion of resistant strains would be higher with the antiretroviral treatment. We should pay more attention to the transmission of resistant strains and continue the drug resistance surveillance.

Acquired Immunodeficiency Syndrome ; virology ; Adolescent ; Adult ; Anti-HIV Agents ; pharmacology ; Child ; Child, Preschool ; China ; Cross-Sectional Studies ; Drug Resistance, Viral ; drug effects ; genetics ; HIV-1 ; drug effects ; genetics ; Humans ; Mutation ; Young Adult

OBJECTIVEDendritic cells play an important role in the pathogenesis of atherosclerosis. To explore the effects of hyperglycemia on the maturation and immune function of human monocyte derived dendritic cells (MDCs).

METHODSImmature MDCs were cultured in RPMI1640 medium with either 5.5 mmol/L D-glucose (NG), 25 mmol/L D-glucose (HG) or 5.5 mmol/L D-glucose + 19.5 mmol/L mannitol (HM) in the absence or presence of 30 mmol/L N-acetylcysteine [NAC, a reactive oxygen species inhibitor (ROS)] for 48 hours. FACS was used to investigate the MDCs immunophenotypic expression. Immune function was evaluated by allogeneic mixed T lymphocyte reaction and measurement of cytokine levels from culture supernatants. Intracellular ROS production in MDCs was also measured by 2', 7'-dichlorodihydrofluorescein (DCF, 10 micromol/L) fluorescence using confocal laser-scanning microscopy techniques.

RESULTSCompared with NG and HM treated MDCs, the expression of maturation markers such as CD1a, HLA-DR, CD83, CD86 were significantly upregulated, allogeneic T cells proliferation as well as the cytokines secretions (IL-2, IL-12, IL-10 and IFN-gamma) significantly increased in HG treated MDCs. Intracellular ROS production in MDCs was also significantly increased and all these stimulatory effects of HG could be partially attenuated by NAC.

CONCLUSIONHigh glucose promote the maturation of MDCs and augment their capacity to stimulate T-cell proliferation and cytokine secretions at least in part through enhancing intracellular ROS generation. These stimulating effects of high glucose on MDCs maturation may be one of the mechanisms of accelerated atherosclerosis found in patients with diabetes.

Cell Differentiation ; drug effects ; Cell Proliferation ; drug effects ; Cells, Cultured ; Culture Media ; Cytokines ; biosynthesis ; Dendritic Cells ; drug effects ; immunology ; metabolism ; Glucose ; adverse effects ; pharmacology ; Humans ; Immunophenotyping ; Monocytes ; cytology ; Reactive Oxygen Species ; metabolism ; T-Lymphocytes ; cytology

Objective:To master the status of Kashin-Beck disease (KBD) in 2019, to provide the basis for assessment of KBD control and elimination.Methods:Data of endemic areas basic information collection and children KBD examination were executed in all endemic villages from every endemic county (city, district, banner) of 13 endemic provinces. All children aged 7 - 12 years in endemic villages underwent clinical examination, and X-ray examination was performed for clinically positive children. When both the clinical examination and X-ray reexamination were positive, the diagnosis was KBD.Results:In monitoring of 823 365 7 - 12 years old children, a total of 3 057 children with similar clinical signs of KBD were suspected positive cases. The results of X-ray reexamination showed that the X-ray manifestations of 3 057 children were normal, and no X-ray positive changes were found, that is, there was no case of KBD in children. A total of 16 559 endemic villages in 13 endemic provinces were monitored, and all reached the criteria for KBD elimination. Surveillance of all endemic villages was completed except Tibet Autonomous Region, the KBD elimination rates of endemic villages were 100.00% in 12 endemic provinces and 99.01% (16 559/16 725) in all 13 endemic provinces.Conclusions:No children KBD case is detected in 2019, children KBD stays at its eliminating level throughout the country. And 100.00% endemic villages meet the criteria for KBD elimination in the remaining 12 endemic provinces except Tibet Autonomous Region.

Objective:To investigate the effects of glucosamine sulfate, chondroitin sulfate and diacetarine on urinary renal function indexes UREA, creatinine (CREA), urinary microprotein(mALB) and N-acetyl-β-D-glucosidase (NAG) in adult patients with Kashin-Beck disease.Methods:According to the criteria of "Diagnosis of Kashin-Beck Disease" (WS/T 207-2010), adult patients with degrees Ⅰ and Ⅱ Kashin-Beck disease in Heilongjiang Province were selected in 2019. They were randomly divided into three treatment groups according to age, gender, disease classification and other condition by clinical randomized controlled trial, group A (glucosamine sulfate group), group B (chondroitin sulfate group) and group C (diacetarine group). Fasting mid-morning urine was collected at 0, 90 and 180 days of treatment. The levels of UREA, CREA, mALB and NAG were measured using an automatic biochemical analyzer. And the abnormal rates of the above indexes were analyzed.Results:At 0 day of treatment, there were 118, 99 and 116 people in the 3 groups, respectively; after 90 days of treatment, 115, 93 and 106 people remained in the 3 groups; after 180 days of treatment, 95, 80 and 93 people remained in the 3 groups. The results showed that there was no significant difference in the levels of UREA, CREA, NAG and mALB among the 3 groups at 0 and 180 days of treatment ( H = 0.055, 0.923, 0.276, 1.125, 1.635, 3.873, 1.045, 4.135, P > 0.05). After 90 days of treatment, there was no significant difference in CREA level among the 3 groups ( H = 1.719, P > 0.05), the levels of UREA and NAG in group C were higher than those in group B ( P < 0.05), and the level of mALB in group B was higher than that in group C ( P < 0.05). The comparison results of all indexes before and after treatment showed that after 90 days of treatment, the levels of mALB in the 3 groups were lower than those of 0 day ( Z = - 2.858, - 3.217, - 2.124, P < 0.05), the levels of NAG were higher than those of 0 day ( Z = - 3.700, - 2.222, - 4.672, P < 0.05); and the level of UREA in group C was higher than that of 0 day ( Z = - 2.393, P < 0.05). After 180 days of treatment, the levels of CREA in the 3 groups were higher than those of 0 day ( Z = - 5.853, - 6.984, - 6.255, P < 0.05), and the levels of mALB in the 3 groups were lower than those of 0 day ( Z = - 3.785, - 2.624, - 3.427, P < 0.05). The abnormal rates of CREA in the 3 groups after 180 days of treatment were higher than those of 0 and 90 days (χ 2 = 39.499, 37.707, 71.534, 57.959, 58.160, 55.129, P < 0.05). There was no significant difference in the abnormal rate of CREA between 0 day and 90 days of treatment (χ 2 = 0.004, 2.068, 0.053, P > 0.05). The abnormal rates of NAG in groups A and C after 90 days of treatment were higher than those of 0 day (χ 2 = 8.999, 11.227, P < 0.05). The abnormal rates of NAG in group C after 180 days of treatment was higher than that of 0 day (χ 2 = 5.006, P < 0.05). There was no significant difference in the abnormal rate of NAG between group A and group C after 90 days and 180 days of treatment (χ 2 = 1.976, 1.413, P > 0.05). The abnormal rates of mALB in groups A and B after 90 days and 180 days of treatment were lower than those of 0 day (χ 2 = 6.461, 8.881, 7.563, 4.999, P < 0.05), and there was no significant difference between 90 days of treatment and 180 days of treatment (χ 2 = 0.638, 0.013, P > 0.05). Conclusions:The effects of glucosamine sulfate, compound chondroitin sulfate and diacetarine on renal function of the patients are not significantly different after 180 days of medication, but the three drugs all have certain effects on CREA and NAG. Follow-up work should be done during drug treatment to closely monitor the changes of the two indicators.