Objective To investigate the regulatory role of the programmed cell death protein 1 (PD-1) and its ligand programmed cell death protein ligand 1 (PD-L1) signaling on the subtypes and functions of natural killer (NK) cells at the maternal-fetal interface during the second trimester in mice following Toxoplasma gondii infection during the first trimester. Methods Twelve 6- to 8-week-old female mice of the C57BL/6J strain were divided into a control group and an infection group, of 6 mice in each group. On the 6.5th day of pregnancy (Gd6.5), each pregnant mouse in the infection group was intraperitoneally injected with 150 tachyzoites of the Toxoplasma gondii PRU strain, while mice in the control group were injected with an equal volume of physiological saline. On the 12.5th day of pregnancy (Gd12.5), uterus and placenta tissues were sampled from pregnant mice for pathological observations, and the mRNA expression levels of PD-1, PD-L1, and tumor necrosis factor-α (TNF-α) were quantified in uterus and placenta tissues. The PD-1 and DX5 expression was measured on NK cells at the maternal-fetal interface using flow cytometry. In addition, the in vitro JEG-3 trophoblast cells and NK-92MI cells co-culture system was established as the control group, and the addition of T. gondii tachyzoites in the co-culture system served as the infection group. The PD-1, PD-L1, and DX5 mRNA expression was quantified in cells using real-time fluorescence quantitative reverse transcription PCR (RT-qPCR) assay, and the TNF-α concentration was measured in the cell culture supernatant using enzyme-linked immunosorbent assay (ELISA). Results On Gd12.5, clear and intact cellular structures of placental decidual tissues were seen in pregnant mice in the control group, with no remarkable abnormal changes found in the uterine columnar epithelial cells, and inflammatory cell infiltration and blood stasis at varying degrees were found in uterine and placental tissues from pregnant mice in the infection group. The relative PD-1, PD-L1, and TNF-α mRNA expression was (1.004 ± 0.004), (1.001 ± 0.001), and (1.001 ± 0.001) in uterine tissues from pregnant mice in the control group and (2.480 ± 0.720), (3.355 ± 0.920), and (2.391 ± 0.073) in the infection group, respectively. The relative PD-1, PD-L1, and TNF-α mRNA expression was (1.007 ± 0.010), (1.006 ± 0.006), and (1.001 ± 0.001) in the uterine tissues in the control group and (6.948 ± 1.918), (3.225 ± 1.034), and (1.536 ± 0.150) in the infection group, respectively. The relative PD-1, PD-L1, and TNF-α mRNA expression was higher in both the uterine (t = 3.55, 4.43 and 33.02, all P values < 0.05) and placental tissues (t = 5.36, 3.72 and 6.18, all P values < 0.05) in the infection group than in the control group. Flow cytometry showed that the proportions of PD-1⁺ NK cells, PD-1⁺ DX5⁺ NK cells, and DX5⁺ NK cells were (12.200 ± 1.082)%, (9.373 ± 7.728)%, and (44.000 ± 4.095)% in uterine tissues from pregnant mice in the control group, and (21.733 ± 1.630)%, (18.767 ± 1.242)%, and (73.367 ± 0.611)% in the infection group, respectively. The proportions of PD-1⁺ NK cells, PD-1⁺ DX5⁺ NK cells, and DX5⁺ NK cells were (1.100 ± 0.510)%, (2.277 ± 1.337)%, and (96.167 ± 2.831)% in placental tissues from mice in the control group, and (26.867 ± 9.722)%, (23.433 ± 6.983)%, and (82.467 ± 2.248)% in the infection group, respectively. The proportions of PD-1⁺ NK cells (t = 8.45, P < 0.05) and DX5⁺ NK cells (t = 12.29, P < 0.05) were higher in uterine tissues from pregnant mice in the infection group than in the control group, and no significant difference was seen in the proportion of PD-1⁺ DX5⁺ NK cells (Z = -1.09, P > 0.05). The proportions of PD-1⁺ NK cells (t = 4.58, P < 0.05) and PD-1⁺ DX5⁺ NK cells (t = 5.15, P < 0.05) were higher in placental tissues from pregnant mice in the infection group than in the control group, while the proportion of DX5⁺ NK cells was lower in the infection group than in the control group (t = -6.56, P < 0.05). RT-qPCR assay revealed that the relative PD-1, PD-L1, and DX5 mRNA expression was (1.010 ± 0.005), (1.002 ± 0.003), and (1.001 ± 0.001) in the JEG-3 cells and NK92MI cells co-culture system and (3.638 ± 1.258), (0.397 ± 0.158), and (4.267 ± 1.750) in the control group, and ELISA measured that the TNF-α concentration was higher in the cell culture supernatant in the infection group [(22.056 ± 3.205) pg/mL] than in the control group [(12.441 ± 0.001) pg/mL] (t = 5.20, P < 0.05). The PD-1(t = 3.62, P < 0.05) and DX5 mRNA expression (t = 3.23, P < 0.05) was higher in the infection group than in the control group, and the PD-L1 mRNA expression was lower in the infection group than in the control group (t = -6.63, P < 0.05). Conclusions Following T. gondii infection, both PD-L1 expression and PD-1 expression on DX5⁺ NK cells at the maternal-fetal interface are upregulated in mice during the second trimester; however, the proportion of DX5⁺ NK cells decreases. These findings suggest that PD-1/PD-L1 signaling may suppress NK cell functions by modulating DX5⁺ NK cell subsets.

Objective To investigate the dynamic expression of cluster of differentiation 47 (CD47) and its ligands signaling regulatory protein α (SIRPα) and thrombospondin-1 (TSP-1) in mice infected with Toxoplasma gondii in the second and third trimesters.. Methods C57BL/6J mice (6 to 8 weeks old) were used for modeling T. gondii infection in the first trimester, and the pregnant mice were randomly divided into the normal control and infection groups, of 10 mice in each group. Pregnant mice in the infection group were intraperitoneally injected with 150 T. gondii tachyzoites on gestational day (Gd) 6.5, while pregnant mice in the normal control group were intraperitoneally injected with the same volume of physiological saline at the same time. The uterine and placental specimens were collected from all pregnant mice on Gd12.5 and Gd18.5, and the pregnant outcomes were recorded. The pathological damages of mouse uterine and placental specimens were observed using hematoxylin-eosin (HE) staining on Gd12.5 and Gd18.5. The relative expression of CD47, SIRPα, TSP-1, surface antigen 1 (SAG1), interferon-γ (IFN-γ), interleukin-2 (IL-2), IL-4 and IL-13 mRNA was quantified in mouse uterine and placental specimens using real-time fluorescence quantitative PCR (qPCR) assay, and the CD47, SIRPα, TSP-1 expression was determined in mouse uterine and placental specimens using immunohistochemical staining. Results As compared with those in the normal control group, the pregnant mice in the infection group showed back arching, bristling, trembling and listlessness during pregnancy, and several mice presented virginal bleeding and abortion. Pathological examinations showed inflammatory cell infiltration, congestion and necrosis in uterine and placental specimens of pregnant mice in the infection group, a higher abortion rate of pregnant mice was seen in the infection group than in the normal control group on Gd12.5 (χ² = 20.405, P < 0.001) and Gd18.5 (χ² = 28.644, P < 0.001). qPCR assay showed significant differences in the expression of CD47, SIRPα, TSP-1, SAG1, INF-γ, IL-2, IL-4 and IL-13 genes in mouse placental specimens between the normal control and infection groups on Gd12.5 and Gd18.5 [F′ (F) = 37.511, 29.337, 97.343, 53.755, 67.188, 21.145, 8.658 and 13.930, all P values < 0.001]. Higher CD47, SIRPα and TSP-1 gene expression was quantified in mouse placental specimens in the infection group than in the normal control group on Gd12.5 (all P values < 0.01), and lower CD47, SIRPα and TSP-1 gene expression was quantified in the infection group than in the normal control group on Gd18.5 (all P values < 0.001), while higher SAG1 gene expression was detected in placental specimens of pregnant mice in the infection group than in the normal control group on Gd12.5 and Gd18.5 (both P values < 0.01). In addition, higher INF-γ and IL-2 expression and lower IL-4 and IL-13 expression was detected in mouse placental specimens in the infection group than in the normal control group on Gd12.5 and Gd18.5 (all P values < 0.001), and there were significant differences in the CD47, SIRPα, TSP-1, SAG1, INF-γ, IL-2, IL-4 and IL-13 gene expression in uterine specimens of pregnant mice between the normal control and infection groups on Gd12.5 and Gd18.5 [H(F′ and F) = 14.951, 25.977, 18.711, 48.595, 39.318, 14.248 and 15.468, all P values < 0.01], and higher CD47 and TSP-1 expression was detected in mouse uterine specimens in the infection group than in the control group on Gd12.5 and Gd18.5 (all P values < 0.01); however, no significant difference was found in the SIRPα expression (P > 0.05). Higher SAG1 expression was detected in uterine specimens of pregnant mice in the infection group than in the normal control group on Gd12.5 and Gd18.5 (both P values < 0.01), and higher INF-γ and IL-2 gene expression and lower IL-4 and IL-13 gene expression was found in the placental specimens of pregnant mice in the infection group than in the normal control group on Gd12.5 and Gd18.5 (all P values < 0.001). Spearman correlation analysis showed that the CD47 gene expression correlated positively with IFN-γ (r_s = 0.735, P < 0.05) and IL-2 (r_s = 0.655, P < 0.05) and negatively with IL-4 (r_s = −0.689, P < 0.05) and IL-13 expression (r_s = −0.795, P < 0.05) in the placental specimens of pregnant mice in the infection group on Gd12.5, and the CD47 gene expression correlated negatively with IFN-γ (r_s = −0.745, P < 0.05) and IL-2 expression (r_s = −0.816, P < 0.05) and positively with IL-4 (r_s = 0.704, P < 0.05) and IL-13 (r_s = 0.802, P < 0.05) in the placental specimens of pregnant mice in the infection group on Gd18.5. Immunohistochemical staining showed mild CD47, SIRPα and TSP-1 expression in uterine and placental specimens of pregnant mice in the normal control group on Gd12.5 and Gd18.5, strong CD47, SIRPα and TSP-1 expression in the placental specimens of pregnant mice in the infection group on Gd12.5 and strong CD47 and TSP-1 expression in the uterine specimens of pregnant mice in the infection group on Gd12.5. Conclusions T. gondii infection in the first trimester may cause abnormal expression of CD47 and its ligands SIRPα and TSP-1 in the maternal-fetal interface of pregnant mice in the second and third trimesters, which may be associated with the immune escape of T. gondii at the maternal-fetal interface.

Objective To predict the structure and antigenic epitope of the Strongyloides stercoralis serine protease inhibitor 1 (Ss-SRPN-1) protein using bioinformatics tools, and to construct prokaryotic expression plasmids for expression of recombinant Ss-SRPN-1 protein, so as to provide the basis for unraveling the function of the Ss-SRPN-1 protein. Methods The amino acid sequence of the Ss-SRPN-1 protein was downloaded from the NCBI database, and the physicochemical properties, structure and antigenic epitopes of the Ss-SRPN-1 protein were predicted using bioinformatics tools, including ExPASy, SWISS-MODEL and Protean. Primers were designed according to the nucleotide sequences of Ss-SRPN-1, and the Ss-SRPN-1 gene was amplified, cloned and sequenced with genomic DNA extracted from the infective third-stage larvae of S. stercoralis as a template. The Ss-SRPN-1 protein sequence was cloned into the pET28a (+) expression vector and transformed into Escherichia coli BL21 (DE) cells for induction of the recombinant Ss-SRPN-1 protein expression. The recombinant Ss-SRPN-1 protein was then purified and identified using Western blotting and mass spectrometry. Results Bioinformatics analysis showed that the Ss-SRPN-1 protein, which was composed of 372 amino acids and had a molecular formula of C₁₉₄₈H₃₀₄₆N₄₈₈O₅₇₅S₁₆, was a stable hydrophilic protein, and the subcellular localization of the protein was predicted to be extracellular. The Ss-SRPN-1 protein was predicted to contain 11 dominant B-cell antigenic epitopes and 20 T-cell antigenic epitopes. The Ss-SRPN-1 gene with a length of 1 119 bp was successfully amplified, and the recombinant plasmid pET28a (+)/Ss-SRPN-1 was constructed and transformed into E. coli BL21(DE) cells. The expressed recombinant Ss-SRPN-1 protein had a molecular weight of approximately 43 kDa, and was characterized as a Ss-SRPN-1 protein. Conclusions The recombinant Ss-SRPN-1 protein has been expressed successfully, and this recombinant protein may be a potential vaccine candidate against strongyloidiasis.

Strongyloides stercoralis is an opportunistic pathogenic parasite that can cause severe strongyloidiasis and even death among immunocompromised individuals. Previous clinical studies have reported cases co-infected with S. stercoralis and other pathogens, such as parasites, viruses, bacteria and fungi. This review summarizes strongyloidiasis patients co-infected with pathogens, and analyzes the impact of co-infection on strongyloidiasis, so as to provide insights into the reduction of the morbidity and mortality of disorders associated with S. stercoralis infections.

In order to fulfill the new requirement of educational and teaching reform in the new era, the Teaching and Research Section of Parasitology in Guangxi Medical University have constructed online courses of Parasites Invasion— Human Parasitology on the platform of Xueyin Online, providing resources in professional learning and general education. In the construction concept, we recombine teaching content to give consideration to both scientificity and interest. In terms of the curriculum content, the curriculum includes micro lecture, PPT, in-lass quiz, case analysis and other diversified teaching resources. In the curriculum management, teachers interact with students in real time with the help of various functions of the platform to fully play the role of supervision. In the assessment, teachers pay attention to students' learning process and carry out comprehensive evaluation. Since the online course was put into operation, good teaching effects has been achieved.

Synthetic polymer hydrogel nanoparticles(NPs)were developed to function as abiotic affinity reagents for fibrinogen.These NPs were made using both temperature-sensitive N-isopropyl acrylamide(NIPAm)and L-amino acid monomers.Five kinds of L-amino acids were acryloylated to obtain functional mono-mers:L-phenylalanine(Phe)and L-leucine(Leu)with hydrophobic side chains,L-glutamic acid(Glu)with negative charges,and L-lysine(Lys)and L-arginine(Arg)with positive charges.After incubating the NPs with fibrinogen,y-globulin,and human serum albumin(HSA)respectively,the NPs that incorporated N-acryloyl-Arg monomers(AArg@NPs)showed the strongest and most specific binding affinity to fibrin-ogen,when compared with y-globulin and HSA.Additionally,the fibrinogen-AArg binding model had the best docking scores,and this may be due to the interaction of positively charged AArg@NPs and the negatively charged fibrinogen D domain and the hydrophobic interaction between them.The specific adsorption of AArg@NPs to fibrinogen was also confirmed by the immunoprecipitation assay,as the AArg@NPs selectively trapped the fibrinogen from a human plasma protein mixture.AArg@NPs had a strong selectivity for,and specificity to,fibrinogen and may be developed as a potential human fibrinogen-specific affinity reagent.

Objective To explore the value of applying flipped classroom in human parasitology. Methods Totally 430 students of 5-year program were divided into experimental group and control group. The experimental class received human parasitology teaching through flipped classroom teaching mode, while the control class received traditional teaching. The effect of teaching was evaluated by questionnaire and examination. The data were analyzed through t-test. Result Meanwhile, statistical difference was found in aver age score of total between experiment group and control group [(68.2 ±8.6) vs. (66.6 ±11.0), P=0.032]. There was also statistical difference in average score of comprehensive analysis [(16.4±3.2) vs. (16.1 ±3.9), P=0.038]. Questionnaire survey of satisfaction showed that 191 students of experimental class (90.95%) felt new teaching mode could improve autonomous learning ability, 199 students (94.76%) in-creased interest in learning;185 students (88.10%) had more interactive with teachers, 178 students (84.76%) enhanced cooperation between st udents, 186 students (88.57%) approved of small group discussion learning and 165 students (78.57%) had no extra burden. Conclusion Flipped classroom teaching mode can improve students' autonomous learning ability and cultivate their abilities of independent thinking, cooperation, criti-cism, innovation, analyzing and solving problems. Thus this new teaching mode is worthy of reference and popularization.

Objective To investigate the effect of acupuncture plus Chinese herbal medication on plasma endothelin (ET-1) and calcitonin gene-related peptide (CGRP) in child patients with mesenteric lymphadenitis.Methods One hundred and eighty child patients with mesenteric lymphadenitis were randomly allocated to groups A, B and C, 60 cases each. Group A received acupuncture at Zusanli and pricking Sifeng points plus oral administration of Wudang Babao Zijinding; group B, oral administration of amoxicillin and clavulanate potassium granules; group C, oral administration of Wudang Babao Zijinding alone. ET-1 and CGRP contents were measured in the three groups before and after treatment.Results There were statistically significant pre-/post-treatment differences in ET-1 and CGRP contents in group A (P<0.01). There were statistically significant post-treatment differences in ET-1 and CGRP contents between group A and group B or C (P<0.01).Conclusions Acupuncture plus Chinese herbal medication is an effective way to treat mesenteric lymphadenitis in children. It can regulate ET-1 and CGRP in the patients.

Objective To analyze the change of cysteinyl leukotriene ( Cys-LTs) levels and 8-Isopros-tane (8-iso-PG) levels in the exhaled breath condensate (EBC) of asthmatic children from acute exacerbation to clinical remission, and investigate the role of the detection of Cys-LTs and 8-iso-PG in EBC in its severity and pathogenesis , and explore the relationship between the Cys-LTs and 8-iso-PG through measuring Cys-LTs levels and 8-iso-PG levels in the EBC of asthmatic children. Methods The outpatient or inpatient asthmatic children of the pediatrics and a group of healthy children were studied. All subjects′ EBC were collected by the R-Tube produced by American Respiratory Research. The concentration of Cys-LTs and 8-iso-PG in EBC were measured by enzyme-linked immunosorbent assay (ELISA), and compared among children in asthmatic exacerbation, asthmatic remission, and healthy condition. The relevance of their change would be explored at the same time. Results (1) Cys-LTs levels in EBC were higher in asthma exacerbation, compared to healthy controls (P<0. 05), while no significant difference were found between asthmatic remission and asthmatic exacerbation or healthy controls ( P>0. 05 ) . ( 2 ) 8-iso-PG levels was higher in asthmatic exacerbation compared to asth-matic remission ( P<0. 05 ) . Moreover, the 8-iso-PG levels were significantly higher in asthmatic remission than in healthy controls (P<0. 05). (3) Through the relevance analysis of the Cys-LTs and 8-iso-PG levels in EBC among the three groups, Cys-LTs levels in EBC of asthmatic exacerbation significantly were correlated with 8-iso-PG levels (n1 =35, r1 =0. 61, P<0. 05), while there was no significant correlation between 8-iso-PG levels and Cys-LTs levels in asthmatic remission. Conclusion The increase of 8-iso-PG levels in EBC of bronchial asthmatic patients correlates with the disease and its control. Therefore, 8-iso-PG can be an objective indicator for asthmatic diagnosis and healing efficacy. Cy-LTs levels increase in the EBC of bronchial asthmatic according to disease severity. The two levels correlate during asthmatic exacerbation, indicating that a link be-tween airway oxidative stress and inflammation among asthmatics.

TLR4-MD-2 complex plays a key role in LPS recognition and its signal transduction. These steps are the vital elements of the host's defensive reaction. Studying the functional domain of TLR4 and MD-2 is very important to further understand the mechanism of LPS signal transduction. It was studied the interaction domain of TLR4 and MD-2 in living cells based on gene mutation, gene transfection and fluorescence resonance energy tramsfer(FRET) which is considered as one of the best methods used for intracellular protein-protein interaction study. CY-15P which was fused by CFP and YFP through 15 neutral amino acids was used as positive control, while co-expressed CFP and YFP proteins were used as negative control. The results showed that the ability of TLR4 binding to MD-2 decreased dramatically after the deletion of Glu~(24) ～ Met~(41) in N terminal of TLR4. Aggregation of TLR4 to LPS stimulation was observed, however, TLR4 without the Glu~(24)～ Met~(41) mutation did not aggregate. All these results indicated that TLR4 Glu~(24)～ Met~(41) might be the interaction domain of TLR4 binding to MD-2 and participate in the aggregation effect of TLR4 upon LPS stimulation.