Objective:To investigate changes in the peripheral interleukin-35 (IL-35) level in patients with alopecia areata, and to assess its modulatory effect on regulatory T (Treg) cell activities.Methods:Totally, 81 patients with alopecia areata (alopecia areata group) and 27 healthy volunteers (control group) were enrolled from Shanxi Provincial People′s Hospital between December 2019 and January 2021. Sera and peripheral blood mononuclear cells (PBMCs) were isolated. Enzyme-linked immunosorbent assay (ELISA) was performed to detect the serum IL-35 level, real-time fluorescence-based quantitative PCR to determine the mRNA expression of IL-35 subunits EBI3 and IL-12p35, and flow cytometry to determine the proportion of CD4 + CD25 + CD127 dim/- Treg cells. Sorted Treg cells were stimulated by recombinant human IL-35, ELISA was performed to detect levels of perforin and granzyme B in the culture supernatant, and real-time fluorescence-based quantitative PCR to determine the mRNA expression of EBI3, IL-12p35, and immune checkpoint molecules, such as programmed death protein 1 (PD-1) , T cell immunoglobulin and mucin protein-3 (Tim-3) , cytotoxic T lymphocyte-associated antigen 4 (CTLA-4) and lymphocyte activation gene-3 (LAG-3) in Treg cells. IL-35-stimulated or unstimulated Treg cells were co-cultured with autologous PBMCs, and cell counting kit-8 (CCK8) assay was conducted to assess cellular proliferative activity. Measurement data were compared between 2 groups by using t test, comparisons among multiple groups were carried out by using one-way analysis of variance, correlation analysis was carried out by using Pearson correlation analysis, and enumeration data were compared by using chi-square test. Results:Compared with the control group, the alopecia areata group showed significantly decreased IL-35 levels (90.10 ± 11.98 ng/L vs. 100.74 ± 28.71 ng/L, t= 2.71, P= 0.008) , mRNA expression of EBI3 and IL-12p35 in PBMCs (EBI3: 1.06 ± 0.15 vs. 1.25 ± 0.11, t= 6.09, P < 0.001; IL-12p35: 1.00 ± 0.15 vs. 1.38 ± 0.22, t= 10.16, P < 0.001) , and proportions of Treg cells (5.91% ± 1.17% vs. 6.85% ± 1.23%, t= 3.54, P= 0.001) . In the alopecia areata group, the proportion of Treg cells was positively correlated with the serum IL-35 level ( r= 0.25, P= 0.026) , and the mRNA expression of EBI3 and IL-12p35 in PBMCs ( r= 0.31, 0.24, P= 0.004, 0.032, respectively) . Compared with the control group, the unstimulated Treg cells from the alopecia areata group showed significantly decreased supernatant levels of perforin and granzyme B, mRNA expression of EBI3, IL-12p35 and immune checkpoint molecules ( P < 0.05 or 0.001) , as well as weakened inhibitory effect on the proliferative activity of PBMCs ( P= 0.013) . There was no significant difference in the level of perforin or granzyme B between the recombinant human IL-35-stimulated and unstimulated Treg cells from the patients with alopecia areata (both P > 0.05) . However, the mRNA expression of EBI3, IL-12p35 and immune checkpoint molecules was significantly higher in the IL-35-stimulated Treg cells than in the unstimulated Treg cells in the alopecia areata group ( P < 0.05 or 0.001) , and the inhibitory effect on the proliferative activity of PBMCs was also significantly enhanced in the IL-35-stimulated Treg cells compared with the unstimulated Treg cells ( P= 0.037) . Conclusion:The peripheral IL-35 level was significantly decreased in the patients with alopecia areata, which was closely associated with reduced activities of Treg cells, and IL-35 may be involved in the occurrence of alopecia areata.

Objective:To investigate changes of natural killer (NK) cell subsets and interleukin-18 (IL-18) level in peripheral blood of patients with alopecia areata, and to assess the regulatory effect of IL-18 on NK cell activity.Methods:A total of 67 patients with alopecia areata (alopecia areata group) and 25 healthy volunteers (control group) were collected from Shanxi Provincial People′s Hospital between December 2019 and January 2021. Peripheral blood mononuclear cells (PBMCs) and plasma were isolated. The percentage of NK cell subsets was investigated by flow cytometry, and plasma IL-18 level was measured by enzyme-linked immunosorbent assay. PBMCs were stimulated with recombinant human IL-18, and co-culture systems of PBMCs with 721.221 cells, K562 cells and P815-Ab cells were established separately. NK cell function was assessed by determining the percentage of CD107a-expressing NK cells and fluorescence intensity of CD16 + NK cells. Comparisons between groups were performed using t test or paired t test. Results:Compared with the control group, the alopecia areata group showed significantly decreased percentage of CD56 +CD16 - NK cells (8.12% ± 3.14% vs. 10.78% ± 4.08%, t = 3.33, P = 0.001) , but significantly increased percentage of CD56 +CD16 + NK cells (46.08% ± 15.21% vs. 32.14% ± 10.45%, t = 4.22, P < 0.001) , and there was no significant difference in the percentage of CD56 -CD16 + NK cells between the alopecia areata group and control group (28.81% ± 8.65% vs. 27.09% ± 7.62%, t = 0.88, P = 0.383) . The plasma IL-18 level was significantly higher in the alopecia areata group than in the control group (112.0 ± 23.72 pg/ml vs. 99.34 ± 15.15 pg/ml, t = 2.48, P = 0.015) . After co-culture with 721.221 cells, K562 cells and P815-Ab cells, the percentage of CD107a-expressing NK cells was significantly higher in NK cells from the alopecia areata group (9.53% ± 1.70%, 5.15% ± 1.35%, 6.50% ± 1.64%, respectively) than in those from the control group (5.00% ± 1.17%, 4.40% ± 1.09%, 5.13% ± 1.36%, respectively, all P < 0.05) . After the stimulation with P815-Ab cells, the alopecia areata group showed significantly decreased fluorescence intensity of CD16 + NK cells (151.10% ± 59.30%) compared with the control group (221.90% ± 93.56%, t = 4.31, P < 0.001) . After IL-18 stimulation, the percentage of CD107a-expressing NK cells significantly increased in the co-culture system of NK cells with 721.221 cells compared with the unstimulated co-culture system (14.47% ± 2.67% vs. 9.93% ± 1.94%, t = 6.00, P < 0.001) , while there was no significant difference between the IL-8-stimulated co-culture system of NK cells with K562 cells or P815-Ab cells and the unstimulated co-culture systems (both P > 0.05) . Conclusion:IL-18 could enhance NK cell activity in patients with alopecia areata, likely by promoting natural cytotoxicity receptor-mediated cytotoxicity.

【Objective】 To investigate the death time of patients with coronavirus disease 2019 (COVID-19). 【Methods】 The death time was calculated and analyzed using individual data and aggregated data through the daily notification of the epidemic situation and the death cases published on the website of the Heath Commission of China and provinces. 【Results】 In the 153 patients who died of COVID-19, the shortest time from onset to death was 4 days and the longest time was 50 days with the mean±standard deviation of (16.7±9.2) days. The median was 14 days and the 95% confidence interval was 4.6-42.9. The shortest time from admission to death was 1 day and the longest time was 50 days with the mean ± standard deviation of (12.1±7.8) days. The median was 11 days and the 95% confidence interval was 2-32.8. The time curve from diagnosis to death was skewed. The death time from diagnosis to death was 0 to 48 days with the mean ± standard deviation of (11.1±8.9) days. The median was 9 days, the interquartile interval was 10.5 days, and the 95% confidence interval was 0-35.4. It took 3 days from onset to admission and 1 day from admission to diagnosis. Aggregated data showed that the time from diagnosis to death of COVID-19 patients in China, China (except Hubei Province), Hubei Province and Wuhan City was 8, 9, 6 and 6 days, respectively. 【Conclusion】 The time from diagnosis to death of COVID-19 patients varied significantly, with the median time of 6-9 days in different regions.

Objective: To assess the role of T cell immunoglobulin and mucin domain-containing protein 3 (TIM-3) in regulation of interleukin (IL) -12 secretion by CD14⁺ peripheral blood monocytes from patients with psoriasis vulgaris. Methods: From November 2017 to March 2018, a total of 47 patients with psoriasis vulgaris (psoriasis group) and 19 healthy volunteers (control group) were enrolled from Shanxi Provincial People′s Hospital. Peripheral blood mononuclear cells (PBMC) were isolated, and stimulated with Toll-like receptor (TLR) ligands. TIM-3 expression and IL-12 secretion by CD14⁺ monocytes were measured by flow cytometry. After blockade of TIM-3 pathway by anti-TIM-3 neutralizing antibody, changes in the downstream signaling pathway molecules in and IL-12 secretion by CD14⁺ monocytes were investigated. Two independent samples t-test was used for comparison between two groups, and Pearson correlation test for correlation analysis. Results: Under the unstimulated condition, the level of IL-12 secreted by CD14⁺ monocytes was very low, and the proportion of CD14⁺ TIM-3⁺ cells was significantly higher in the psoriasis group (12.20% ± 2.83%) than in the control group (9.91% ± 1.77%, t = 3.270, P = 0.001 7) . After the stimulation with TLR ligands, the proportion of CD14⁺ IL-12⁺ cells was significantly lower in the psoriasis group (13.49% ± 2.80%) than in the control group (28.97% ± 8.97%, t = 10.71, P < 0.000 1) , but the proportion of CD14⁺TIM-3⁺ cells was still higher in the psoriasis group (6.80% ± 1.11%) than in the control group (4.85% ± 1.37%, t = 6.064, P < 0.000 1) . The proportion of CD14⁺TIM-3⁺ cells was negatively correlated with that of CD14⁺ IL-12⁺ cells in both the control group and psoriasis group (r = -0.473, -0.371 respectively, both P < 0.05) . The TIM-3 pathway blockade could induce the decrease of suppressor of cytokine signaling 1 in CD14⁺ monocytes, increase the phosphorylation of signaling transducers and activators of transcription 1, and promote IL-12 secretion by CD14⁺ monocytes induced by keratinocytes isolated from psoriatic skin lesions. Conclusion TIM-3 plays a crucial role in the negative regulation of CD14⁺ monocyte-induced innate immune response in psoriasis.

Objective To investigate the clinical effectiveness of extended care on coronary intervention therapy in patients with acute myocardial infarction. Methods 90 patients with acute myocardial infarction who had been through coronary intervention therapy successfully in our hospital from June 2014 to April 2015 were divided into study group and control group with 45 patients in each according random number table. Patients in control group were treated with routine nursing care intervention while patients in study group were treated with additional extended care. Clinical effectiveness of nursing care in two groups were observed. Results After the intervention of extended care, there were 2 cases with nonfatal myocardial infarction (4. 4％), 3 patients undergoing revascularization for a second time (6. 7％) and one death (2. 2％) in study group, which were significantly fewer than those in control group, the difference was statistically significant(P＜0. 05). 30 cases were satisfied with extended care and 11 cases were somewhat satisfied in study group after intervention. The satisfactory rate in study group was 91. 1％, which was significantly higher than those in control group (66. 7％), the difference was statistically significant (P＜0. 05). According to the results of follow-up visit, nursing compliance of diet, exercise, medication and review in patients of study group was better than that in control group, the differences were statistically significant(P＜0. 05). Conclusion Extended care on coronary intervention therapy in patients with acute myocardial infarction could reduce the incidence of adverse reaction, improve the satisfactory rate of nursing care and improve patients ' quality of life. It was worth promotion.

Objective To evaluate masticatory performance and life quality of children with ectodermal dysplasia(ED) after prosthetic rehabilitation.Methods Six children with ED received denture restoration and 18 healthy children were involved in this study.The surface electromyography(EMG) of masseter(MM) and anterior temporalis(TA) during clenching and chewing movement were recorded.The EMG amplitude,area,asymmetry index of total and activity index of MM/TA were compared at each stage.The masticatory efficiency was measured with spectrophotometer.The life quality was assessed using visual analogue scale questionnaire.Results The EMG amplitude of MM and TA during chewing in ED Group were 41.7％ and 45.6％ of the control group respectively,the area were 35.9％ and 36.0％ respectively.Significant difference in asymmetry index of total during clenching was observed between the two groups (P＜0.05) but not during chewing(P＞0.05).The differences of activity index of MM/TA during clenching and chewing between the two groups were not detected(P＞0.05).The masticatory efficiency of ED group was 67.2％ of the control group.The score of chewing function in children with ED after prosthetic rehabilitation was three times higher than before,and no difference was present between the two groups (P＞0.05).Conclusions Early prosthetic rehabilitation can significantly improve the masticatory performance and life quality of children with ED.

Objective To explore the effects of aberrant expression of DNA methyltransferase 1 (DNMT1) in cervical cancerization tissue and cervical cancer cells.Methods Cervical tissues were collected from 80 cases with a diagnosis of invasive cervix squamous cell carcinoma (SCC),53 cases with high-grade cervical intraepithelial neoplasia (CIN Ⅱ/Ⅲ),52 cases with low-grade cervical intraepithelial neoplasia (CIN Ⅰ)and 53 cases with normal cervix (NC).Meanwhile,Caski (HPV16-positive) and C33A (HPV-negative) cells selected from cervical cancer cell lines were cultured routinely in vitro.The expression of DNMT1 protein and mRNA were examined by Western blot analysis and real-time PCR (qRT-PCR) in the tissues and cells,respectively.Results The levels of DNMT1 protein were 1.33,1.84 and 2.28,and the Ct-ratios (DNMT1/β-actin) of DNMT1 mRNA were 1.27,1.27 and 1.26 in CIN Ⅰ,CIN Ⅱ/Ⅲand SCC group,respectively.Comparing with NC group,the expression of DNMT1 protein or mRNA was elevated in deficient cervical groups,with statistical significance (F =110.57,P ＜ 0.001,F =2.68,P =0.048).The expression levels of DNMT1 protein were increased steadily according to severity of the cervix lesions (x2tend =50.80,P ＜ 0.001),however,the expression of DNMT1 mRNA was not observed the same tendency (x2tend =3.63,P ＞ 0.05).The results from experiment in vitro showed that the levels of DNMT1 protein or mRNA were both higher in Caski cell than in C33A cell,especially for DNMT1 mRNA with significantly difference (t =7.134,P =0.002).Conclusion Aberrant expression of DNMT1 protein or mRNA could link with the risk of cervical cancerization by both transcriptional and posttranscriptional mechanisms.There would be a synergistic effect between overexpression of DNMT1 and HPV16 infection in the progression of cervix carcinogenesis.

Objective To analyze risk factors for metabolic syndrome in Mongolia versus Han nationalities in Xilinhaote city. Methods Using the epidemiology investigation data of health examination,we calculated the prevalence of metabolic syndrome of Mongolia and Han nationalities, then used logistic regression model to explore risk factors of two nationalities. Results The crude prevalence of MS in Mongolia and Han nationality was 34.3%and 24.6% respectively. The multivariate logistic regression showed that male, the meat-rich diets and aging(OR:2.18, 1.92, 1.04 respectively)were the risk factors for Mongolia nationality, and smoking, family history of hypertension, drinking, the meat-rich diets, aging(OR:1.89, 1.84, 1.72, 1.61 and 1.04 respectively)were the risk factors for Han nationality. Conclusions Xinlinhaote population has higher MS prevalence, and different nationalities have different risk factors. We should take preventive actions to control it.

AIM: To investigate the feasibility of establishment for chronic graft-versus-host disease (cGVHD) model in rats after allo-bone marrow transplantation (allo-BMT), in order to provide premise conditions for further studying the immuno-regulation role of mesenchymal stem cell (MSC) on GVHD after allo-BMT. METHODS: This experiment was finished in Laboratory of Pathology and Pathophysiology in Sun Yat-sen University from April to September 2006.①Six-week-old male Fischer344 rats (RT1Al) were used as donors while six-week-old female Waster (RT1Au) rats were used as recipients.②Recipient rats were given water supplemented with gentamycin (320 mg/L) and erythromycin (250 mg/L) three days before BMT. On the day of transplantation, recipient rats received 8.0 Gy (60Co ?, 0.7 Gy/min) total body irradiation. Within 6 hours following the irradiation, recipient rats in BMT group were transplanted with 0.8?108 cells via tail vein injection, while rats in control group only received the same volume injection of phosphate buffer. Each group included 10 animals. Evaluation of common living status was monitored including daily diet, activity, stool and urine, fur and body mass. Shaved skin, liver and intestine tissues were also analyzed histologically. RESULTS: ①All rats in the control group died within 17 days following the irradiation and most of them died on day 11 or 12 post-transplantation, while BMT group had higher survival rate, in addition to three rats died on days 12, 16, 18 respectively, whereas others were all alive through 60 days expectation period.②Rats in the BMT group had no clear symptoms of acute GVHD, such as rapid weight loss and severe diarrhea, however, the weight growth in rats of the BMT group was quite slow. Furthermore, 1 month following BMT, depilation phenomenon was evident in the head and back of recipients, and then extended to abdominal part and extremity with the increase of time. Two months following BMT, skin follicular dropout and slight dermal mononuclear infiltration were found. Hepatic disease was characterized by portal tract lymphocyte infiltration, fibrous thickening and sclerosis of the bile duct wall. Small bowel specimens showed clear inflammatory cell infiltration (neutrophils, acidophils, macrophages) within lamina propria. CONCLUSION: ①The cGVHD model can be established through allo-BMT from F344 to Wistar rats.②The typical histological signs of cGVHD are evident in skin, liver and intestine tissues, among which hepatic sign is the most dominant including portal tract and bile duct mononuclear infiltration followed by fibrous thickening and sclerosis of the bile duct wall.

We have constructed the immune microsphere against tumor necrosis factor-alpha (TNF-alpha) prospectively, hoping to establish the experiment groundwork in more researches which could be used in specific elimination of the TNF-alpha by blood purification method for the future. The recombinant human tumor necrosis factor-alpha monoclonal antibody (rHTNF-alpha McAb) was wrapped on the polystyrene microsphere (PSM) carrier connecting poly-L-lysine (PLL) beforehand. They were earmarked by the fluorescein isothiocyanate (FITC) respectively. The packing conditions were examined using the inversted and fluorescence microscopes and the spectrophotometer. The results showed that the best conditions for wrapping were 20 degrees C, pH9.5 and 60 minutes. The PLL content was not changed in the washing fluid after coating, which indicated the wrapping was quite firm. At the same temperature and same coating time, the rHTNF-alpha McAb coated on the PLL was obviously substantial when the concentration of glutaraldehyde solution was 0.2%. The findings demonstrated that the built immune microsphers can be used as a novel adsorption material. This method is simple and economic, and it offers a new approach to the related studies.
Antibodies, Monoclonal ; immunology ; Fluorescein ; chemistry ; Humans ; Microspheres ; Polylysine ; chemistry ; Polystyrenes ; chemistry ; Recombinant Proteins ; immunology ; Tumor Necrosis Factor-alpha ; immunology