Yuejie Zhu

TGF-β/Smad signaling pathway regulates Th17/Treg balance during Echinococcus multilocularis infection

Alveolar echinococcosis (AE) is a severe parasitic disease caused by the infection of Echinococcus multilocularis (Em). Very little is known on the relationship between TGF-β/Smad signaling pathway and Treg/Th17 balance in the infected liver at different periods after Em infection. Using qRT-PCR, immunohistochemistry, flow cytometry and CBA assay, we measured the expression levels of TGF-β, Smad2/3/7, ROR-γt, Foxp3, IL-17, IL-10 and percentages of Th17 cells and Treg cells in mouse AE model, from day 2 to day 270 after infection. In the early stage of infection (day 2 to day 30), Smad7 was up-regulated and the TGF-β pathway was inactivated. In the middle stage of infection (day 30 to day 90), TGF-β and Smad2/3 were up-regulated. And levels of Treg cells, Foxp3, Th17 cells, RORγt, IL-17, IL-10 and IL-6 were significantly increased. In the late stage of infection (day 90 to day 270), Treg cells, Foxp3, TGF-β and IL-10 maintained at high levels whereas Th17 cells and IL-17 decreased significantly. TGF-β/Smad signaling pathway was activated during the chronic infection. Our data suggest that there were Treg/Th17 imbalance in the middle and especially in the late stage of Em infection and that Treg/Th17 imbalance may be regulated by TGF-β/Smad signaling pathway. Treg and Th17 subsets may be involved in regulating immune tolerance and tissue inflammation, and facilitating the long-term survival of Em in the host.

The Th17/Treg balance and the expression of related cytokines in Uygur cervical cancer patients

BackgroundThe fine balance of Th17/Treg is crucial for maintenance of immune homeostasis. The objective of this study was to investigate the balance of Th17/Treg and the expression of related cytokines in Uighur cervical cancer patients.MethodsPeripheral blood was collected from 65 cases of cervical cancer patients, 42 cases of cervical CIN patients and 40 healthy people. Flow cytometry was used to detect the percentages of T cell subsets, including CD3+ T cells, CD4+ T cells, CD8+ T cells, Treg cells and Th17 cells. ELISA assay was conducted to detect expression levels of TGF-β, IL-6, IL-10, IL-17, IL-23 and IFN-γ.ResultsThere were no significant difference in the levels of CD3+ T cells, CD4+ T cells, CD8+ T cells, and the ratio of CD4+/CD8+ among the cervical cancer group, the CIN group and the healthy control group. However, compared with the healthy control group, the percentages of CD4+ CD25+ Treg, CD4+CD25+CD127- Treg, CD4+IL17+ Th17, CD4+CD25+Foxp3+, CD4+CD25- Foxp3+, CD8+CD25+CD127-Treg and CD8+CD25+Foxp3 were significantly higher in the cervical cancer group and the CIN group. Similar results were also found in the Th17/Treg ratio and the related cytokines. There was no significant difference between the cervical cancer group and the CIN group. Additionally, Th17 cell levels were positively correlated with IL-6, IL-23 and IL-17. Also, Treg cell levels were positively correlated with TGF-β, IL-10 and IL-6. Contrarily, Treg cell levels and IFN-γ were negatively correlated.ConclusionsOur data indicated that the Th17/Treg balance was broken in peripheral blood of cervical cancer patients. Analysis of Th17/Treg balance may have a significant implication in diagnosing cervical cancer.Virtual slidesThe virtual slide for this article can be found here: http://www.diagnosticpathology.diagnomx.eu/vs/1813823795931511

Th9/IL-9 Profile in Human Echinococcosis: Their Involvement in Immune Response during Infection by Echinococcus granulosus

Th9 cells have been reported to contribute to immune responses; however, the role of Th9 cells in Echinococcus granulosus infection is unknown. This study is to determine whether Th9 cells and IL-9 are involved in human Echinococcus granulosus infection. Compared with healthy controls (HC group), the mRNA levels of PU.1, IL-9, and GATA-3 were significantly increased in patients before therapy (CE group), as revealed by qRT-PCR. Flow cytometry analysis showed that the percentages of Th9 and Th2 cells in CE group were significantly higher. The levels of IL-9, IL-4, IL-10, and TGF-β in CE group were also significantly increased, as detected by CBA assay. The percentages of Th9 and Th2 cells in CE group were positively correlated. After treatments of surgery in combination with albendazole, the PU.1 and GATA-3 mRNA levels were significantly decreased in patients after therapy (PCE group) compared with CE group. The numbers of Th9 and Th2 cells and levels of IL-9, IL-4, IL-10, and TGF-β were also significantly decreased in PCE group. In conclusion, the ratios of Th9 cells and IL-9 levels were significantly decreased after treatment, suggesting that Th9/IL-9 may be involved in immune response induced by Echinococcus granulosus infection.

Genotype distribution and the relative risk factors for human papillomavirus in Urumqi, China

The aim of this study was to investigate human papillomavirus (HPV) infection and HPV genotype distributions in Urumqi, Xinjiang, China. The related risk factors for high-risk HPV infection was also analyzed. A stratified cluster sampling method was used for the population-based cervical cancer screening of women aged 18–69 years in the Urumqi Saybagh district. Exfoliated cervical cell samples were collected for liquid-based cytology detection and HPV genotyping DNA microarrays. Education level, number of sexual partners, condom use and occupation were used in the multivariate analysis model. The HPV infection rate of women working in service industries was significantly higher compared with those of white-collar workers, community residents and migrant workers. The 35–44-year-old migrant worker group had the highest HPV infection rates among all of the groups in the three different age ranges. The number of marriages, education level, smoking history, number of abortions, use of condoms, number of sexual partners, number of sexual partners in the past five years and occupation were all associated with female HPV infection rate (P<0.05). The 35–44-year-old women were the age group with the highest HPV infection rate. The HPV infection rate of females in service industries was the highest. Education level and condom use were protective factors of HPV infection, while the number of sexual partners and occupation were risk factors for HPV infection.

The prediction of T‑ and B‑combined epitope and tertiary structure of the Eg95 antigen of Echinococcus granulosus

Echinococcosis, also known as hydatid disease, is a type of zoonotic parasitic disease caused by the Echinococcus larvae infection. The disease is severely harmful to both humans and animals. Research and development of an epitope vaccine is crucial. To determine the dominant epitopes of the Eg95 antigen, the tertiary structure and the T- and B-combined epitope of the Eg95 protein for Echinococcus granulosus were predicted and analyzed in the present study. The tertiary structure of the Eg95 protein was predicted using the 3DLigandsite server and RasMol software. The T- and B-combined epitope of the Eg95 antigen was analyzed using the DNAStar (V5.0), IEDB, SYFPEITHI and BIMAS. Tertiary structure prediction results showed that there were potential epitopes in Eg95 antigen. Bioinformatics analysis revealed the T- and B-combined epitopes of Eg95 antigen. Four and six T- and B-combined epitopes induced immune responses in humans and mice. Additionally, four T- and B-combined epitopes induced immune responses in both humans and mice. The tertiary structure and T- and B-combined epitopes of the Eg95 protein were also determined. The results obtained in the present study may be beneficial in the investigation of Eg95 antigenicity and the development of dominant epitope vaccines.

Elevated Expression of Programmed Death-1 and Programmed Death Ligand-1 Negatively Regulates Immune Response against Cervical Cancer Cells

The present study is to measure the expression of programmed death-1 (PD-1) and programmed death ligand-1 (PD-L1), as well as its clinical significance in cervical cancer patients. Our results showed that different T cell subsets in patients with cervical cancer had high expression of PD-1, and DCs had high expression of PD-L1. High expression of PD-1 on Treg cells in cervical cancer patients facilitated the production of TGF-β and IL-10 but inhibited the production of IFN-γ. Cervical cancer elevated the expression of PD-1 and PD-L1 in mRNA level. PD-1 expression in peripheral blood of cervical cancer patients was related with tumor differentiation, lymph node metastasis, and invasiveness. PD-1/PD-L1 pathway inhibited lymphocyte proliferation but enhanced the secretion of IL-10 and TGF-β in vitro. In summary, our findings demonstrate that elevated levels of PD-1/PD-L1, TGF-β, and IL-10 in peripheral blood of cervical cancer patients may negatively regulate immune response against cervical cancer cells and contribute to the progression of cervical cancer. Therefore, PD-1/PD-L1 pathway may become an immunotherapy target in the future.

CCR7(lo)PD-1(hi) CXCR5(+) CD4(+) T cells are positively correlated with levels of IL-21 in active and transitional cystic echinococcosis patients.

BackgroundIn our study, we investigated whether circulating T follicular helper (Tfh) and the related cytokines are involved in human cystic echinococcosis (CE).MethodsA total of 64 patients with CE and 30 healthy controls were enrolled in this study. Percentages of CCR7loPD-1hi cells within CXCR5+ CD4+ T cells (circulating Tfh cells) were detected by flow cytometry. Levels of IL-21 and IL-4 in peripheral blood were detected by cytometric bead array. The mRNA expression of IL-21, IL-4, Bcl-6, and Blimp-1 in peripheral blood mononuclear cells (PBMCs) were measured by real-time PCR. Levels of IgG1, IgG2, IgG3, and IgG4 in the patients’ sera were measured using enzyme-linked immunosorbent assay.ResultsPercentages of circulating Tfh cells were significantly increased in the CE1, CE2, and CE3 groups (p < 0.05). The concentrations of IL-21 and IL-4 in the serum were significantly increased in CE1, CE2, and CE3 groups (p < 0.05). IL-21 was positively correlated with circulating Tfh cells in CE3 group (r = 0.779, p < 0.05). The mRNA levels of IL-21, IL-4, and Bcl-6 were increased in CE1, CE2, and CE3 groups. Levels of IgG1 and IgG4 in patients’ sera were increased in CE1, CE2, and CE3 groups. Levels of IgG2 and IgG3 were increased in CE4-5 group. Additionally, after stimulation with hydatid fluid in vitro, the levels of circulating Tfh cells, IL-21 and IL-4 in PBMCs isolated from CE patients were significantly increased (p < 0.05).ConclusionsThe levels of circulating Tfh and related cytokines were significantly increased in CE patients, suggesting that they are involved in human CE.

Activity in mice of recombinant BCG-EgG1Y162 vaccine for Echinococcus granulosus infection

Cystic hydatid disease is a zoonotic parasitic disease caused by Echinococcus granulosus which is distributed worldwide. The disease is difficult to treat with surgery removal is the only cure treatment. In the high endemic areas, vaccination of humans is believed a way to protect communities from the disease. In this study we vaccinated BALB/c mice with rBCG-EgG1Y162, and then detected the level of IgG and IgE specifically against the recombinant protein by ELISA, rBCG-EgG1Y162 induced strong and specific cellular and humoral immune responses. In vitro study showed that rBCG-EgG1Y162 vaccine not only promote splenocytes proliferation but also active T cell. In addition, the rBCG-EgG1Y162 induced a protection in the mice against secondary infection of Echinococcus granulosus.

Increased levels of CCR7(lo)PD‑1(hi) CXCR5+ CD4+ T cells, and associated factors Bcl‑6, CXCR5, IL‑21 and IL‑6 contribute to repeated implantation failure

In vitro fertilization-embryo transfer (IVF-ET) can be used by infertile couples to assist with reproduction; however, failure of the embryo to implant into the endometrial lining results in failure of the IVF treatment. The present study investigated the expression of chemokine receptor 7 (CCR7)(lo) programmed death-1(PD-1)(hi) chemokine receptor type 5 (CXCR5)+ cluster of differentiation 4 (CD4)+ T cells and associated factors in patients with repeated implantation failure (RIF). A total of 30 females with RIF and 30 healthy females were enrolled in the current study. Flow cytometry was used to detect the proportion of CCR7(lo)PD-1(hi) CXCR5+ CD4+ T cells in the peripheral blood. Cytokine bead arrays were performed to detect the levels of interleukin (IL)-6, −4 and −2 in the serum. ELISAs were used to detect the level of IL-21 in the serum. Quantitative real time polymerase chain reaction analysis and immunohistochemistry were used to investigate the expression of B-cell lymphoma 6 (Bcl-6), chemokine receptor type 5 (CXCR5) and IL-21 in the endometrium. The results revealed that the percentage of CCR7(lo)PD-1(hi) CXCR5+ CD4+ T cells was increased in the RIF group compared with the control group during the mid luteal phase. The mRNA and protein levels of Bcl-6, IL-21 and CXCR5 in the endometrium and the concentrations of IL-21 and IL-6 in the serum were significantly increased in the RIF group; however, no significant difference was observed between the two groups in regards to the expression of IL-4 and IL-2. Furthermore, a significant positive correlation was identified between the percentage of CCR7(lo)PD-1(hi) CXCR5+ CD4+ T cells and IL-21 and IL-6 levels. The expression of IL-21 also had a positive correlation with Bcl-6 and CXCR5 expression in the RIF group. These results suggest that increased levels of CCR7(lo)PD-1(hi) CXCR5+ CD4+ T cells and associated factors contribute to RIF and could therefore be a potential therapeutic target.

Immunization of mice with egG1Y162-1/2 provides protection against Echinococcus granulosus infection in BALB/c mice

Aims: This study is to predict and purify the T‐B combined epitopes of egG1Y162 antigen in Echinococcus granulosus, and to evaluate their immunogenicity in mice. Methods: The bioinformatics software was used to predict the T‐B combined epitopes of egG1Y162 antigen. Recombinant egG1Y161/2 peptides were constructed, expressed and purified. Mice were immunized with egG1Y161/2 peptides. The serum and spleen cells were isolated. The isolated spleen cells were stimulated with egG1Y161/2 peptides in vitro and the culture supernatant was collected. The levels of IgG in serum and levels of IL‐4 and IFN‐&ggr; in the culture supernatant were measured by ELISA. The weight and number of the fresh hydatid cysts were evaluated. The serum ptotoscolicidal activity was measured by the complement dependent cytotoxicity assay. Results: Peptides of 6–19aa, 64–82aa, 106–119aa were predicted as T‐B combined epitopes of egG1Y162 antigen. And, recombinant protein egG1Y162–1 or egG1Y162–2, which contained T‐B combined epitope(s) of the 6–19aa, or the 64–82aa and the 106–119aa in egG1Y162 antigen, respectively, was successfully expressed and purified. Serum IgG levels of mice immunized with egG1Y162–1/2 were significantly increased during the immune response to Echinococcus granulosus. The levels of IFN‐&ggr;, IL‐4 and the ratio of IFN‐&ggr;/IL‐4 after egG1Y162–1/2 immunization were significantly higher. Weight and number of the fresh hydatid cysts in egG1Y162–1/2 immunized mice was significantly decreased. And, the serum protoscolicidal activity after egG1Y162–1/2 immunization was enhanced. Conclusions: The egG1Y162–1/2 induces production of serum IgG levels and Th1 cell immune response, which enhances the protective immunity in Echinococcus granulosus challenged mice and thus may be used as a potential vaccine candidate. HIGHLIGHTST‐B combined epitopes of egG1Y162 antigen are predicted.EgG1Y162–1/2 protein is expressed and purified in vitro.The IgG levels in serum are increased after EgG1Y162–1/2 immunization.egG1Y162–1/2 enhances protective immunity in mice.