Objective To investigate the protective effect of chloroquine on endotoxemia mice and its inhibition on the release of cytokines induced by LPS. Methods A total of 40 mice of Kunming species were randomly divided into four groups: LPS group received LPS at 10 mg/kg, chloroquine group received chloroquine at 20 mg/kg, LPS plus chloroquine group received chloroquine at 20 mg/kg first, then LPS at 10 mg/kg and control group received only 0.9%sodium chloride at 200 ?l/20 g. The mortality was observed within seven days after injection via caudal vein. ANA 1 cell lines were cultivated in vitro . After chloroquine was first added into the cells for 3 hours, the releases of TNF ? and IL 6 in the supernatants induced by different concentrations of LPS were measured. Results Chloroquine could decrease the death of mice due to endotoxin. Mortality dropped from 100% to 50% ( P

Objective To construct an Apoptin prokaryotic vector,aiming to produce antigenic fusion protein Apoptin. Methods The Apoptin gene was amplified from the template of plasmid pSSCHG/NT4-Apoptin-HA2-TAT by PCR.The Apoptin was sub-cloned into the multiple clone sites of plasmid pET-28a(+) to get the prokaryotic vector of pET-28a(+)-Apoptin,which was transformed into E.coli BL21(DE3).Expression of E.coli BL21(DE3) was induced by IPTG.The specific protein expression was detected by SDS-PAGE. Results The fusion protein was expressed with high efficiency in E.coli BL21(DE3) transformed by pET-28a(+)-Apoptin after induction with IPTG.The specific fusion protein had an apparent related molecular weight of about 17 000 ku as indicated by SDA-PAGE analysis. Conclusion The Apoptin prokaryotic expression vector with pET-28a(+)-Apoptin can effectively express Apoptin fusion protein,laying a foundation for further study of Apoptin and preparation of antibodies against Apoptin.

OBJECTIVEIn vitro model of hydrogen peroxide induced apoptosis of SW-480 cells was used to investigate the role of NF-kappaB in the pathogenesis of reactive oxygen species induced apoptosis of intestinal epithelial cells.

METHODSUltra-structural changes were observed. Apoptosis of SW-480 cell line was determined by Annexin-V and PI double-stained flow cytometry. Nuclear translocation of NF-kappaB was determined by anti-NF-kappaB polyclonal antibody and EB double-staining. NF-kappaB activity was studied by electrophoretic mobility shift assays. RT-PCR was performed to study expression of NF-kappaB mRNA.

RESULTSHydrogen peroxide led to apoptosis of SW-480 cells, condensed or semilunar chromatin even apoptotic bodies could be observed. Nuclear translocation of NF-kappaB, increase of NF-kappaB activity and expression of NF-kappaB mRNA were found simultaneously.

CONCLUSIONSEarly activation of NF-kappaB may be one of the mechanisms of apoptosis in intestinal epithelial cells by reactive oxygen species.

Apoptosis ; Humans ; Hydrogen Peroxide ; toxicity ; Intestinal Mucosa ; metabolism ; Microscopy, Confocal ; NF-kappa B ; metabolism ; Reactive Oxygen Species ; toxicity ; Tumor Cells, Cultured

Objective To study the effect of pioglitazone on the differentiation and function of rat osteoclast-like cells (OLC), and to probe the relationship between activated PPARγ2 and osteoclasts. Methods On day 1 of OLC formation from nonadherent bone marrow ceils (BMC) obtained from rats induced by M-CSF and receptor activator of NF-кB ligand (RANKL), 1, 5 and 10μmol/L pioglitazone hydrochloride was added. RT- PCR was performed to determine the mRNA expressions of PPARγ2 and receptor activator of NF-кB (RANK) on day 3, 5 and 7 during incubation, the number of tartrate-resistant acid phosphatase (TRAP)-positive cells,the number of bone resorption pits and the ratio of its area on dentin slice were counted, the activity of TRAP and the mean fluorescence intensity of integrin β3 (CD61) of OLC were also measured. Results (1) The effect on the differentiation of OLC: The addition of pioglitazone at the start of the culture period induced a dose-dependent decrease in TRAP-positive OLC and the activity of TRAP (P < 0.01 or P < 0.05) ; the mRNA expression of PPARγ2 was up-regulated by 5 and 10 μmol/L pioglitazone in the early stage of incubation and attenuated with thematuration of OLC on the contrary, however, the expression of RANK was down-regulated by 5 and 10 μmol/L piolitazone in every stage of incubation (P < 0.05 or P < 0.01), combined with decrease in TRAP-positive OLC from day 3 by 10 μmol/L pioglitazone. (2) The effect on the function of OLC: the number of bone resorption pits and the ratio of its area on dentin slice were decreased in groups of 5 and 10 μmol/L pioglitazone (P < 0.01 orP < 0.05), no obvious change was noted in the group with 1 μmol/L pioglitazone compared with the control group; the mean fluorescence intensity of CD61 were down-regulated in groups of 5 and 10 μmol/L pioglitazone (P < 0.05 or P <0.01). Conclusion Activation of PPARγ2 pathway by pioglitazone could partially inhibit differentiation and function of OLC derived from rat BMC.

OBJECTIVETo observe the role of four modeling peptides (10342, 10343, 10344, 10345) derived from bactericidal/permeability increasing protein (BPI) in neutralizing endotoxin (LPS) in vitro and in vivo.

METHODSQuantitative limulus amoebocyte lysate assay was employed to evaluate the capacity of BPI peptides in neutralizing endotoxin in vitro. The protective capacity of the peptides was observed in mice challenged with endotoxin by intravenous administration via the tail vein. The influence of the peptides on serum TNFalpha and IL-6 levels in rats with endotoxemia were also observed.

RESULTSAll of the four peptides possessed endotoxin-neutralizing capacity which was strengthened along with the increase in their concentration. Among the peptides, 10342 is the strongest one. All of the peptides had strong power to protect mice from endotoxin with 90% protective rate. In the rats with endotoxemia, the four peptides could reduce the levels of serum TNFalpha and IL-6 significantly at different time-points.

CONCLUSIONFour BPI modeling peptides possessed not only endotoxin-neutralizing capacity in vitro, but also potential protective capacity in animals challenged with endotoxin.

Animals ; Anti-Infective Agents ; immunology ; pharmacology ; Antimicrobial Cationic Peptides ; Blood Proteins ; immunology ; pharmacology ; Endotoxemia ; metabolism ; prevention & control ; Female ; Interleukin-6 ; blood ; Lipopolysaccharides ; administration & dosage ; immunology ; Male ; Membrane Proteins ; Mice ; Neutralization Tests ; Rats ; Rats, Wistar ; Time Factors ; Tumor Necrosis Factor-alpha ; metabolism

OBJECTIVETo explore the activation of polymorphonuclear cells (PMNs) and vascular endothelial cells co-cultured with and stimulated by lipopolysaccharide.

METHODSPMNs in concentration of 2 x 10(6)/ml isolated from healthy volunteers by Percoll gradient were added to monolayer of ECV-304 cells grown to confluency, then different groups were prepared according to final concentration of lipopolysaccharide. The morphological change was observed under invert microscope. The changes in TNFalpha and IL-6 levels of the supernatant of the cultured cells were determined at 4, 8, 12 and 24 hours after culturation.

RESULTSThe TNFalpha production of cultured pure ECV-304 exhibited no remarkable change when stimulated by different concentrations of LPS. But the TNFalpha production of the ECV-304 increased significantly when co-cultured with PMNs at 4 hr and stimulated by LPS in concentration of 10 micro g/ml, and increased at 8 hours and lasted up to 24 hours of culturation in higher levels (P < 0.05). The IL-6 production of cultured pure ECV-304 increased obviously along with the increase of LPS concentration, and it showed no change when PMNs co-cultured with ECV-304. While the IL-6 level in the supernatant of co-cultured ECV-304 with PMNs increased sharply when stimulated by both low (100 ng/ml) and high (1 micro g/ml) concentrations of LPS and maintained at high levels up to 24 hours of culturation. The higher the concentration of LPS was, the quicker the IL-6 level increased.

CONCLUSIONCo-cultured PMNs and endothelial cells could be activated and activation state could be maitained by low concentrations of LPS.

Adult ; Cell Line ; Cells, Cultured ; Coculture Techniques ; Dose-Response Relationship, Drug ; Endothelium, Vascular ; cytology ; drug effects ; metabolism ; Humans ; Interleukin-6 ; metabolism ; Lipopolysaccharides ; pharmacology ; Neutrophils ; cytology ; drug effects ; metabolism ; Time Factors ; Tumor Necrosis Factor-alpha ; metabolism

Objective:To investigate the correlation between exhaled nitric oxide and obstructive sleep apnea-hypopnea syndrome (OSAHS).Methods:Eighty patients with OSAHS (OSAHS group) who received treatment in Pingxiang People's Hospital from September 2019 to September 2021 were included in this study. An additional 60 patients with snoring (snoring group) who concurrently received treatment in the same hospital were included in the control group. The value of exhaled nitric oxide was measured using an exhaled nitric oxide detector. The relationship between exhaled nitric oxide and apnea-hypopnea index, and the lowest oxygen saturation level during sleep (LSaO 2) was analyzed using Pearson correlation analysis. The optimal cut-off value of exhaled nitric oxide for predicting OSAHS was analyzed using the receiver operating characteristic curve. Results:Exhaled nitric oxide and apnea-hypopnea index in the OSAHS group were (18.61 ± 6.23) μg/L and (44.50 ± 16.15) times/hour, respectively, which were significantly greater than (11.17 ± 4.31) μg/L and (2.91 ± 0.79) times/hour in the snoring group ( t = 7.94, 14.08, both P < 0.05). LSaO 2 in the OSAHS group was significantly lower than that in the snoring group [(66.53 ± 10.17)% vs. (92.15 ± 1.62)%, t = -13.61, P < 0.05]. Correlation analysis showed that exhaled nitric oxide levels in patients with OSAHS were positively correlated with apnea-hyponea index ( r = 0.56, P = 0.001), and negatively correlated with the lowest oxygen saturation level ( r = -0.54, P = 0.002). The receiver operating characteristic curve analysis revealed that when the optimal cut-off value of exhaled nitric oxide was 11.5 μg/L, the area under the curve was 0.846, with sensitivity of 91.3%, and specificity of 63.3%. Conclusion:Patients with OSAHS have airway inflammatory reactions. The level of nitric oxide in exhaled air is positively correlated with the severity of OSAHS, which has a certain clinical value.

Objective To compare the cumulative analgesic effects of electroacupuncture at Sanyinjiao (SP6), Xuanzhong (GB39) and non-acupoint in treating primary dysmenorrhea. Method By adopting a multi-centered randomized controlled study method, 501 patients recruited from Dongzhimen Hospital of Beijing University of Chinese Medicine, China-Japan Friendship Hospital, Beijing Hospital of Traditional Chinese Medicine of Capital Medical University, First Teaching Hospital of Tianjin University of Traditional Chinese Medicine, Huguosi Hospital of Chinese Medicine of Beijing University of Chinese Medicine and the Outpatient of Shandong University of Traditional Chinese Medicine were randomized into a Sanyinjiao group, a Xuanzhong group, and a non-acupoint group, 167 subjects in each group. The electroacupuncture intervention was applied when dysmenorrhea flared up and the Visual Analogue Scale (VAS) ≥40 mm, with frequency at 2/100 Hz and intensity during patient’s endurance, 30 min each time, once a day, and for successive 3 d. Before the first treatment, 30 min after the first treatment, and respectively prior to the second and third treatment, VAS was used to measure the pain intensity. Meanwhile, the Retrospective Symptom Scale (RSS-COX 2) was investigated before the first treatment, right after the removal of needles for the first treatment, before the second and third treatment. Result The decrease of VAS in Sanyinjiao group was more significant than that in Xuanzhong group and non-acupoint group (MD=﹣2.92 mm, P=0.028; MD=﹣3.47 mm, P=0.009), while there was no significant difference between Xuanzhong group and non-acupoint group (MD=﹣0.56 mm, P=0.674); there were no significant differences in comparing the RSS-COX2 total score among the three groups (P=0.086). Conclusion Sanyinjiao (SP6) can produce a more significant cumulative analgesic effect for primary dysmenorrhea patient than Xuanzhong and non-acupoint, and the effects of Xuanzhong and non-acupoit are equivalent.

Objective: To explore the effect of pulsatile lavage on wound healing in diabetic foot ulcer patients. Methods: The random number table method was used to divide 86 patients of diabetic foot ulcers into two groups with 43 patients in each group. The control group disinfected and cleaned the wound by routine methods, while the experimental group received closed pulse irrigation with sewage collection unit. The two groups were debridement, dressing selection and wound dressing in a unified way. The frequency of dressing change, time of dressing change, efficacy, cost of dressing changes, score of wound pain and wound healing were observed. Results: The frequency of dressing change, dressing change time, wound healing time and total effective rate of the experimental group were (10.42±1.92) times, (12.19±2.37) min, (32.53±6.91) d and 86.04% (37/43), respectively, while those of the control group were (19.47±3.13) times, (21.65±3.99) min, (43.17±13.72) d and 51.16% (22/43), with statistically significant differences (t values were 4.545-16.127, χ² value was 13.214, all P < 0.01). However, the cost of dressing change in the experimental group was (3 278.78±220.92) yuan, and that in the control group was (3 195.75±206.54) yuan. There was no significant difference between the two groups (t value was -1.814, P > 0.05). The pain scores were (1.47±1.09), (0.57±0.72), (0.06±0.23), (0.003±0.01) points in the experimental group at the 2nd week,the 4th week,the 6th week and the 8th week after intervention, and they were (3.83±1.16), (2.73±1.41), (1.92±1.06), (1.43±0.70) points respectively in the control group, the differences were statistically significant (F_time=390.663, F_intergroup=76.011, F_interaction=4.210, all P < 0.01). The wound healing rates were (49.34±9.34)%, (86.26±13.33)%, (95.01±8.56)%, (97.28±3.62)% respectively in the experimental group in the 2nd week,the 4th week,the 6th week and the 8th week after intervention,while in the control group they were (26.64±5.19)%, (50.37±10.53)%, (64.84±12.27)% and (72.04±12.96)%. The differences between the two groups were statistically significant (F_time=354.487, F_intergroup= 921.230, F_interaction =23.154, all P < 0.01). Conclusions Pulsatile lavage can effectively clean the wound, reduce the frequency of dressing change, shorten the time of dressing change and wound healing, reduce the wound pain, improve the wound healing rate of diabetic foot ulcers, and did not increase the economic burden of patients, which was worthy of clinical application.