Expression of the immune checkpoint molecules CD226 and TIGIT in preeclampsia patients.

BACKGROUND: Imbalanced immune responses are involved in developing preeclampsia (PE). We wish to explore the expression and potential changes of immune checkpoint molecules TIGIT, CD226 and CD155 in PE patients. METHODS: The expression of the immune checkpoint molecules TIGIT, CD226 and CD155 in different lymphocyte subpopulations was determined by flow cytometry in 24 patients with PE and compared to 24 healthy pregnant women of the same gestational age as the controls.​Serum CD155 was detected by ELISA in the patients with PE compared to controls. RESULTS: The percentages of CD4+ and CD8+ T lymphocytes in the peripheral blood of PE patients were not significantly different from those of the controls, whereas the regulatory T cells (Tregs) in PE patients were significantly lower than those in controls (6.43 ± 1.77% vs. 7.48 ± 1.71%, P = 0.0420). The expression of TIGIT and CD226 showed different percentages on CD4+ T cells, CD8+ T cells and Treg cells. However, the difference in the percentages of TIGIT, CD226 on these T cells between the two groups was not statistically significant. The level of CD155 in peripheral serum of PE patients was 6.64 ± 1.79 ng/ml, which was not significantly different from that in the control group 5.61 ± 1.77 ng/ml, P = 0.0505. The present results demonstrate that TIGIT, CD226 and CD155 are not present at altered immune conditions in the peripheral blood of patients with PE, compared with normal pregnant women. CONCLUSION: The immune checkpoint molecules TIGIT, CD226 and CD155 are not abnormally expressed in PE patients.

Mechanistic insights into aniline-induced liver injury: Role of the mmu_circ_26984/Myh9/NLRP3 axis and modulation by N-acetylcysteine.

Aniline is a widely used chemical. Chronic or high-dose exposure to aniline can lead to hepatocellular damage. Although the hepatic pathogenicity of aniline has been established in previous studies, studies involving pathogenic genes during aniline-induced liver injury are limited. Our study first discovered and identified the role and mechanism underlying a new circRNA mmu_circ_26984 in aniline-induced chemical liver injury. Further, we discuss the protective effect of N-acetylcysteine (NAC) in this pathway. After constructing in vitro and in vivo models of aniline treatment, we screened the circRNA with significant differences in expression in AML12 cells from control and aniline-treated groups by circRNA microarray analysis. Next, using RNA pulldown, liquid chromatography-mass spectrometry (LC-MS), and RNA immunoprecipitation, we analyzed the relationship between mmu_circ_26984 and myosin heavy chain 9 (Myh9). Subsequently, we determined the specific mechanism of action of mmu_circ_26984 and Myh9 in aniline-induced liver injury and the protective effect of NAC against aniline-induced liver injury process using Cell Counting Kit-8, Western blot, RNA extraction, a reverse transcription quantitative polymerase chain reaction (RT-qPCR), fluorescence in situ hybridization, immunohistochemistry, and immunofluorescence. The expression of mmu_circ_26984 was significantly increased in liver tissues and AML12 cells of aniline-treated mice compared with the control group. This high expression of mmu_circ_26984 increased the expression of injury-related inflammatory factors, such as NLRP3, Caspase-1, IL-18, and IL-1ß in vivo and ex vivo, which exacerbated the level of liver injury. The interaction of mmu_circ_26984 with Myh9 also affected the course of liver injury. Mmu_circ_26984 overexpression and reduced treatment affected the levels of Myh9 expression in AML12 cells, as well as downstream inflammatory factors associated with injury, such as NLRP3. In addition, NAC reduced the process of liver injury mediated by the mmu_circ_26984/Myh9/NLRP3 axis. In conclusion, mmu_circ_26984 is a potential molecular marker and therapeutic target in the process of aniline-induced liver injury that can mediate aniline-exposure-induced liver injury via modulation of the mmu_circ_26984/Myh9/NLRP3 axis, and NAC can effectively attenuate the effect of this liver injury.

Aniline exposure activates receptor-interacting serine/threonineprotein kinase 1 and causes necroptosis of AML12 cells.

With the increased use of aniline, potential impacts on human health cannot be ignored. The hepatotoxicity of aniline is largely unknown and the underlying mechanism remains unclear. Therefore, the aim of the present study was to investigate the hepatotoxicity of aniline and elucidate the underlying mechanism. AML12 cells were exposed to different concentrations of aniline (0, 5, 10, or 20 mM) to observe changes to reactive oxygen species (ROS) production and the expression patterns of necroptosis-related proteins (RIPK1, RIPK3, and MLKL). The potential mechanism underlying aniline-induced hepatotoxicity was explored by knockout of RIPK1. The results showed that aniline induced cytotoxicity in AML12 cells in a dose-dependent manner in addition to the production of ROS and subsequent necroptosis of AML12 cells. Silencing of RIPK1 reversed upregulation of necroptosis-related proteins in AML12 cells exposed to aniline, demonstrating that aniline-induced ROS production was related to necroptosis of AML12. Moreover, aniline promoted intracellular RIPK1 activation, suggesting that the RIPK1/ROS pathway plays an important role in aniline-induced hepatotoxicity. NAC could quench ROS and inhibit necroptosis. These results provide a scientific basis for future studies of aniline-induced hepatotoxicity for the prevention and treatment of aniline-induced cytotoxicity.

Molecular epidemiology and risk factors of Stenotrophomonas maltophilia infections in a Chinese teaching hospital.

BACKGROUND: Stenotrophomonas maltophilia (S. maltophilia) is an important opportunistic pathogen that can be isolated in hospitals. With the abuse of broad spectrum antibiotics and invasive surgical devices, the rate of S. maltophilia infection is increasing every year. This study was an epidemiological analysis of the clinical and molecular characteristics of S. maltophilia infection in a Chinese teaching hospital. The goal was to obtain a comprehensive understanding of the status of S. maltophilia infection to provide strong epidemiological data for the prevention and treatment of S. maltophilia infection. RESULTS: A total of 93 isolates from Renji Hospital affiliated with the Shanghai Jiaotong University School of Medicine were included, in which 62 isolates were from male patients. In addition, 81 isolates were isolated from sputum samples. A total of 86 patients had underlying diseases. All patients received antibiotics. Multilocus sequence typing (MLST) analysis indicated that 61 different sequence types (STs) were found (including 45 novel STs), and MLST did not show significantly dominant STs. Pulsed field gel electrophoresis (PFGE) results showed that 93 isolates could be divided into 73 clusters, and they also showed weak genetic linkages between isolates. The resistant rates to trimethoprim/sulfamethoxazole (TMP/SMX) and levofloxacin were 9.7 and 4.3%, respectively, and all isolates were susceptible to minocycline. Four virulence gene's loci Stmpr1, Stmpr2, Smf-1, and Smlt3773 were positive in 79.6, 91.4, 94.6, and 52.7% of the isolates, respectively. Three biofilm genes rmlA, spgM, and rpfF were positive in 82.8, 92.5, and 64.5% of the isolates, respectively. Mean biofilm forming level of OD492 was 0.54 ± 0.49. We did not find any significant difference between different genders and different age-groups. We retrospectively analyzed data from patients in the intensive care unit (ICU) and the control group. The independent risk factors of those who were infected in the ICU included immunosuppression and the increased antibiotic usage. CONCLUSIONS: Most of the patients had prior medical usage histories and baseline diseases. The positive rate of virulence genes was high, the drug resistance rate of S. maltophilia was low, and the biofilm formation ability was strong. The increased use of antibiotics was an independent risk factor for S. maltophilia infection, which should receive more attention. No obvious clonal transmissions were found in the same departments.

Upregulation of the lncRNA SRLR in polycystic ovary syndrome regulates cell apoptosis and IL-6 expression.

Increased levels of interleukin-6 (IL-6) contribute to the development of polycystic ovary syndrome (PCOS); in renal cell carcinoma, the long non-coding RNA (lncRNA) SRLR upregulates IL-6. In this study, we demonstrated that the levels of the lncRNA SRLR were upregulated in PCOS patients with high expression of plasma IL-6 compared with heathy females. The levels of the lncRNA SRLR in the plasma had a positive correlation with expression of IL-6 in patients with PCOS but not in healthy females. Upregulation of the lncRNA SRLR in plasma could distinguish PCOS patients from healthy females. Overexpression of the lncRNA SRLR led to upregulation of IL-6 and promoted apoptosis of human granulosa-like tumour cells (KGN). Therefore, the lncRNA SRLR participated in PCOS by regulating cell apoptosis and IL-6 expression. SIGNIFICANCE OF THE STUDY: The lncRNA SRLR mediates its effects on apoptosis and IL-6 expression in PCOS and could be used to distinguish PCOS patients from healthy controls. Plasma circulating levels of the lncRNA SRLR may be a potential target for the treatment of PCOS.

Multiplex PCR for sexing Schistosoma japonicum cercariae and its utility.

Accurate discrimination of the Schistosoma japonicum cercariae gender is very important for establishing monosexual infection animal models and for standardizing the real intensity of infection. In this study, a multiplex PCR technique consisting of two pairs of primers, of which one amplifies a 185-bp band specific for the W chromosome and the other amplifies a 420-bp band for the Z chromosome, was established to sex the S. japonicum cercariae. For male cercariae (ZZ), a single 420-bp band is expected, and for female cercariea (ZW), two distinct 185-bp and 420-bp bands can be observed. There was no cross-reaction with S. mansoni, S. haematobium, Clonorchis sinensis, Paragonimus westermani, and Trichinella spiralis. After sexing the cercariae escaped from a single snail, mice in group A were infected with 60 male cercariae and mice of group B were infected with 40 female cercariae. Meanwhile, mice in group C were infected with 10 male and 10 female cercariae that were sexed by multiplex PCR. At 45 days postinfection, male and female adult worms were recovered to verify the accuracy of multiplex PCR for sexing S. japonicum cercariae and to calculate the male and female survival rate and paired worm ratio. Our results showed that the multiplex PCR technique could distinguish male cercariae with 100% accuracy. However, sometimes the discrimination results of multiplex PCR mis-scored mixed sexual cercariae as female cercariae. The mean male adult worm burden in mice of group C was 10.7 ± 2.4, and the mean female adult worm burden was 7.7 ± 2.5. There was a significant difference between the male worm burden and female worm burden in group C. The P value was 0.013. The real paired worm ratio of group C was 74.2% (95%CI 56.6~91.8%). These results demonstrated a male-biased sex ratio in the mice model with equilibrated sex ratio cercariae infection, as predicted by our multiplex PCR technique. In conclusion, our multiplex PCR technique is an effective tool for sexing S. japonicum cercariae, especially for distinguishing male cercariae, which is of great value for establishing monosexual cercariae infection mice models to harvest male adult worms for anti-schistosomal drug screening.

Estrogen promotes the survival of human secretory phase endometrial stromal cells via CXCL12/CXCR4 up-regulation-mediated autophagy inhibition.

STUDY QUESTION: What mechanism is involved in regulating the autophagy of endometrial stromal cells (ESCs), and does it participate in the pathogenesis of endometriosis? SUMMARY ANSWER: CXCL12 down-regulates secretory phase ESC autophagy. WHAT IS KNOWN ALREADY: mTOR (mammalian target of rapamycin), the major negative regulator of autophagy, is abnormally increased in endometriotic lesions and is involved in the direct regulation of endometrial stromal cell (ESC) apoptosis. STUDY DESIGN, SIZE, DURATION: Autophagy was measured by transmission electron microscopy and immunofluorescence, and in vitro analysis was used to measure estrogen/CXCL12/CXCR4 signaling-mediated ESC autophagy. PARTICIPANTS/MATERIALS, SETTING, METHODS: A total of 31 controls and 31 women with histologically confirmed endometriosis were included. We measured the autophagy level of normal and endometriosis-derived endometrium, and its relationship to the stage of endometriosis, as well as the potential molecular and signaling pathways that mediate the aberrant autophagy in endometriosis. MAIN RESULTS AND THE ROLE OF CHANCE: Compared with control secretory phase ESCs, a significant reduction of the autophagy grade (as observed in TEM), punctuate LC3B staining (as observed in immunofluorescence assays), and autophagy-associated protein levels were exhibited in secretory phase eutopic ESCs (P < 0.05) and ectopic ESCs (P < 0.05) from women with endometriosis. In addition, the autophagy level was strongly negatively correlated with the CXCL12 concentration in ESCs (R(2) = -0.9694). However, there was no significant difference in autophagy grade or CXCL12 concentration between stage I-II and stage III-IV endometriosis-derived ectopic ESCs (P > 0.05). Based on a human autophagy PCR array, CXCL12 and CXCR4, which is the CXCL12 receptor, in ESCs were predicted to be molecules that mediate the abnormally lower autophagy in endometriosis. Accordingly, after estradiol (E2) treatment a marked increase in CXCL12 secretion (1.71-fold, P < 0.01) and CXCR4 expression (5.07-fold, P < 0.01) in secretory phase ESCs was observed together with decreases in autophagy grade (TEM), punctuate LC3B immunofluorescent staining and autophagy-associated protein levels (P < 0.05). These changes could be reversed by progesterone (P4) (P < 0.05). The suppression of autophagy induced by E2 and recombinant human CXCL12 protein could be abrogated by an anti-CXCR4 neutralizing antibody and by a NF-κB inhibitor (P < 0.05), respectively. In addition, estrogen-stimulated CXCL12 secretion led to a low population of S phase cells (P < 0.05), as well as a low level of apoptosis (P < 0.05) in secretory phase ESCs. LIMITATIONS, REASONS FOR CAUTION: Further studies are needed to examine the mechanism of autophagy on ESC apoptosis. WIDER IMPLICATIONS OF THE FINDINGS: Measures to increase in endometrial autophagy might be a valid, novel approach to reduce local E2-dependent growth of endometriotic tissue. STUDY FUNDING/COMPETING INTERESTS: This study was supported by the National Natural Science Foundation of China (NSFC) (81471513, 81471548 and 81270677), the Training Program for Young Talents of Shanghai Health System XYQ2013104, the Program for Zhuoxue of Fudan University, and the Program for Creative Talents Education of Key Disciplines of Fudan University. None of the authors has any conflict of interest to declare.

Diagnostic value of dual-fluorescence staining in bacterial vaginosis.

OBJECTIVE: The aim of this study was to investigate the epidemiology of bacterial vaginosis (BV) in Shanghai, China, and to explore the value of a dual-fluorescence staining method in the diagnosis of BV. METHODS: Specimens were collected from women with vaginitis at the Obstetrics and Gynecology Hospital of Fudan University from January 2020 to December 2021, and the proportions of various vaginitis types (such as Candida vaginitis, Trichomonas, and bacterial vaginitis) were analyzed statistically. To explore the diagnostic value of dual-fluorescence staining for BV, we first executed a dual-fluorescence staining method to analyze the vaginal secretions of 265 patients, then confirmed our diagnoses by consulting clinical physicians and by using Nugent scoring of Gram staining. RESULTS: There were 16,905 patients who were diagnosed with vaginitis over the previous 2 years, with a median age of 32 (minimum age of 9 years and maximum of 84 years). Of these patients, we noted 10,887 cases (64.40%) of BV. Our staining results revealed that the dual-fluorescence method was consistent with Gram staining in the diagnosis of BV, with a P value of less than .001 using a χ 2 test and a consistency kappa value of 0.896. Compared with Gram staining, the dual-fluorescence staining method required an acceptable time (2.2 min vs 2.5 min, respectively) and exhibited different visual effects (green and yellow vs purple and red, respectively). CONCLUSION: Dual-fluorescence staining for the detection of bacterial diseases of the vagina exhibited acceptable consistency with Gram staining and performed well with respect to dyeing time, stability, and the interpretation of results. We argue that this method should be used in outpatient services.

Decrease in CD226 expression on CD4+ T cells in patients with endometriosis.

Endometriosis is a chronic inflammatory disease. The immune-checkpoint molecules CD226 and TIGIT play an important role in regulating T cells' function. However, little is known about the proportion and function of CD226 and TIGIT on CD4+ T cells in endometriosis. The current study found no significant differences in the TIGIT percentage on peripheral CD4+ T cells between patients with endometriosis and the control group. However, CD226 was lower in patients with endometriosis than that in the control group (P < 0.01). The cytokines TNF-α, IL10, and IFN-γ were significantly elevated in TIGIT+ CD4+ T cells compared to TIGIT- CD4+ T cells. HLA-DR+ cells were more numerous among TIGIT+ CD4+ T cells than among the TIGIT- subset (P <0.001). Similarly, the cytokines TNF-α, IL10, and IFN-γ were significantly elevated in CD226+ CD4+ T cells compared to levels in CD226- CD4+ T cells. The proportion of HLA-DR+ CD4+ T cells among CD226+ CD4+ T cells was also significantly higher than that among the CD226- subset (P < 0.001). After TIGIT was blocked, the level of IL-10 in TIGIT+ CD4+ T cells was higher than that in cells with unblocked TIGIT. There were no differences in TNF-α and IFN-γ. After CD226 was blocked, TNF-α and IFN-γwere lower while IL-10 was higher. In conclusion, there is a diminution of CD226 in CD4+ T cells in patients with endometriosis. This is correlated with the effector function of CD4+ T cells, and blocking CD226 can suppress this function.

Clinical and Molecular Epidemiology of Stenotrophomonas maltophilia in Pediatric Patients From a Chinese Teaching Hospital.

Objective: To study the molecular epidemiological characteristics of Stenotrophomonas maltophilia (SMA) isolated from patients in a pediatric teaching hospital in Shanghai so as to provide data for the prevention and treatment of SMA. Methods: Non-repetitive SMA strains were isolated from patients from January 2013 to December 2014. The cloning characteristics were analyzed using multilocus sequence typing (MLST) and pulsed field gel electrophoresis (PFGE), and the drug resistance was determined using the Kirby-Bauer disk method. Virulence genes and biofilm genes were detected using polymerase chain reaction (PCR). The biofilm forming ability was analyzed using the semi-quantitative biofilm formation test. Results: A total of 104 strains were collected, primarily from the pediatric intensive care unit and thoracic surgery, and these strains were isolated from sputum sources (n = 82). A majority of the patients were male (67/104), and the age range was between 6 days and 12 years old. A total of 95 patients had 1-3 baseline diseases. All of the patients had prior use of 1-4 antimicrobial agents. A total of 59 STs were detected using the MLST analysis, of which 45 were new. The sequence types of the SMA were scattered, with no trend in the clonal spread. The PFGE showed that the 104 strains could be divided into 93 clusters, with no obvious cluster aggregations. All of the strains were susceptible to levofloxacin, trimethoprim/sulfamethoxazole, and minocycline. The positive rates of the virulence genes stmPr1, stmPr2, smf-1, and smlt3773 locus were 98.1, 86.5, 100, and 91.3%, respectively. All of the strains had biofilm formation, and most of the strains had strong biofilm formation abilities. The positive rates of the three biofilm genes rmlA, spgM, and rpfF were 83.7, 100, and 45.2%, respectively. However, the point mutations of rmlA and spgM with strong biofilm formation abilities were significantly different from those with weak biofilm formation abilities. Conclusion: Most infected patients had prior use of antibiotics and underlying diseases, and the positive rate of the virulence gene was high. The strains were susceptible to three kinds of antibiotics and had strong biofilm formation abilities. The mutations of rmlA and spgM may be related to the biofilm formation ability, and no obvious clonal transmissions were found in the same clinical department.

Alterations of Several Serum Parameters Are Associated with Preeclampsia and May Be Potential Markers for the Assessment of PE Severity.

The precise pathophysiological mechanisms of preeclampsia (PE) and preventative strategies remain unknown. Laboratory markers which can help in identifying PE patients from pregnant women and assessing the severity of PE during pregnancy are worthy to be explored. In this study, a retrospective case-control study was designed to assess whether the serum levels of albumin (ALB), total protein (TP), prealbumin (PA), alkaline phosphatase (ALP), lactic dehydrogenase (LDH), D-dimer, fibrinogen (Fbg), platelet (PLT) count, mean platelet volume (MPV), and platelet distribution width (PDW) can help in assessing PE and evaluate its severity. 256 pregnant women were enrolled and classified into 3 groups: mild preeclampsia (mPE, n = 85), severe preeclampsia (sPE, n = 78), and healthy normotensive controls (control, n = 93). Our result showed that the serum levels of ALP, LDH, and D-dimer were significantly higher in mild or severe PE patients compared with the healthy controls (66 (52.5-76.5) vs. 168 (141.5-201.25) vs. 182.5 (120-191.5), 152 (139.75-166.25) vs. 183.5 (163.25-307) vs. 282 (215.25-306), 1.05 (0.65-1.57) vs. 3.05 (2.25-4.08) vs. 5.65 (2.29-7.71)), while ALB, TP, and PA are lower (38 (37-42) vs. 31.5 (25.5-34.5) vs. 28.5 (24-33), 65 (63-68.25) vs. 56.5 (52-61) vs. 51.5 (49-58), 219.14 ± 68.25 vs. 167.88 ± 52.21 vs. 143.22 ± 50.46). On the other hand, compared with the mPE group, the sPE group showed significantly lower PLT count but higher level of LDH, D-dimer, and Fbg. No significant differences in MPV or PDW were found between any of the two groups. In conclusion, the above markers except for the MPV and PDW may be correlated with PE severity in this patient cohort, indicating possible values of these potential biomarkers in auxiliary diagnosis and severity assessment of PE.

The immune dysfunction in ankylosing spondylitis patients.

Ankylosing spondylitis (AS) is a spinal arthritic disease that is often associated with human leukocyte antigen (HLA)-B27, while only part of HLA-B27 carriers become AS patients. T cells have been reported to play an important role in the pathology of AS. T-cell immunoglobulin and mucin-domain-containing molecule 3 (Tim-3) and programmed death-1 (PD-1) have been known to negatively regulate the immune response. In this study, we used flow cytometry to analyze the immunological differences of peripheral bloodfrom 21 patients with AS, 22 cases who didn't have AS but were found to be HLA-B27 positive (HLA-B27+ group), and 16 normal healthy individuals (Healthy group). The level of CD4+, CD8+ T cells,and Treg of each group was observed. The expression of Tim-3 and PD-1 and the production of IFN-γ, IL-6, TNF-α, IL-4, and IL-10 were examined as well. We found that the percentage of Treg in AS group was lower than that of healthy group. The expression of PD-1 on CD8+ T cells and Tim-3 on CD4+ T cells was lower in the AS group. AS group had lower IL-10 production by CD4+ T cells and higher IL-6 production by CD8+ T cells. The results of HLA-B27+ group were similar to that of the healthy group. These data suggested that patients with AS had an impairment in the ability to negatively regulate the immune response, which might be related to the etiology of AS. To further investigate the roles of Tim-3 and PD-1 on is a dysfunction of T cells in AS that is associated with PD-1 and Tim-3.

Up-regulation of Interleukin-21 Contributes to Liver Pathology of Schistosomiasis by Driving GC Immune Responses and Activating HSCs in Mice.

The pathology of schistosome egg-induced liver granuloma, fibrosis and eventually liver scarring is complicated. CD4+ helper T (Th) cells play critical roles in both host humoral immunity and cellular immunity against parasitic infection and immunopathology in schistosomiasis. Follicular helper T (Tfh) cells are another specialized subset of Th cells and involved in infectious diseases. However, the immune regulatory mechanism of Tfh cells in severe liver pathology of schistosomiasis is still poorly understood. In this study, using a S. japonicum-infected mouse model, we studied the dynamics and effects of Tfh cells in vivo and demonstrated that Tfh phenotype molecules ICOS, PD-1 and functional factor IL-21 were positively correlated with disease development by flow cytometry. Meanwhile, our results also showed that Tfh cells enriched in splenic germinal center (GC) and promoted B cells producing IgM with the progress of hepatic immunopathology by B-T co-culture experiments. More importantly, our data indicated that IL-21 contributed to the formation and development of hepatic egg granuloma and subsequent fibrosis by driving GC responses and activating HSCs by immunohistochemical detection and blocking assay in vitro. Our findings contribute to the better understanding of the immunopathogenesis of schistosomiasis and have implications for therapeutic intervention of hepatic fibrotic diseases.

Dynamics of Th9 cells and their potential role in immunopathogenesis of murine schistosomiasis.

BACKGROUND: Th1, Th2, Th17, Treg and Tfh cells play important roles in schistosomiasis. Th9 cells secrete IL-9 as a signature cytokine and contribute to several classes of inflammatory disease. However, the effects of Th9 cells in schistosomiasis are unknown. We aimed to explore the dynamic changes and potential roles of Th9 cells in the pathogenesis of hepatic egg granulomatous inflammation in mice infected with Schistosoma japonicum. METHODS: Twenty mice with S. japonicum infection and five normal controls (NC) were used as models. The average areas of egg granulomas were estimated by hematoxylin-eosin (H & E) staining. Hepatic IL-9 and transcription factor PU.1 levels were detected by immunohistochemistry. Flow cytometry techniques were used to analyze the proportions of Th9 cells. With the help of ELISA, serum levels of IL-9 were examined. RESULTS: The egg granulomas began to form from four weeks after infection and continued to develop. In parallel with the development of egg granulomas, the hepatic levels of IL-9 and PU.1 increased very slowly during the first four weeks post-infection and increased rapidly thereafter. Moreover, the proportions of splenic Th9 cells and levels of serum IL-9 had similar developmental trends with the egg granulomas. CONCLUSION: The proliferation of Th9 cells and levels of IL-9 were significantly higher in S. japonicum-infected mice compared to NC. In addition, dynamic changes of Th9 and IL-9 were synchronous with the developmental trend of hepatic egg granulomatous inflammation, suggesting that Th9 cells might be a new subset in the pathogenesis of schistosomiasis.

Multifarious effects of 17-ß-estradiol on apolipoprotein E receptors gene expression during osteoblast differentiation in vitro.

Apolipoprotein E (ApoE) regulated bone metabolism in mice might mediate uptake of lipid particles into target cells such as osteoblasts via receptor-mediated endocytosis by apoE receptors, which includes the low-density lipoprotein receptor (LDLR) family and heparan sulfate proteoglycans (HSPGs). There is no report regarding the expression of ApoE receptors mRNA induced by estrogen during osteoblast differentiation in vitro. Primary osteoblasts were collected from the calvaria of newborn mice and were subjected to osteoblast mineralization culture with serial concentrations of 17-ß-estradiol (E2) in vitro. RNA was isolated at days 0, 5 and 25 of differentiation. Real-time PCR was conducted to analyze apoE receptors mRNA levels. We found that most LDLR family members genes were induced during osteoblast differentiation in vitro. The effect of E2 on apoE receptors gene expression during osteoblast differentiation was multifarious. The most noted members of the LDLR family involved in the maintenance of bone metabolism were LRP5, LRP6, LRP4, and Apoer2. LRP6 was up-regulated, while LRP5, LRP4, and Apoer2 were down-regulated by E2. Given that LRP6 is required for early stages of differentiation, we speculate E2 promotes osteoblast differentiation mainly in the early stage.

Combination of CD4(+)CD25(+)CD127(-) regulatory T cells with MLC-BE and BE-Ab2: an efficient evaluation of the therapy of paternal lymphocyte induced immunization in unexplained recurrent spontaneous abortion patients.

The aim of this retrospective study was to compare the immune tolerance status of patients suffered from unexplained spontaneous abortion (URSA) before and after treatment with paternal lymphocyte induced immunization (PLII) four times, and its relationship to the pregnancy outcome. 168 URSA patients were included in the present study. Among 168 couples, 138 couples were conceived again, of whom 86 were successfully pregnant till 20 gestational weeks, 31 cases again failed in the first trimester, 21 cases were still under follow-up, another 30 cases still had not conceived. Both the level of one way mixed lymphocyte culture blocking efficiency (MLC-BE) and anti-idio blocking antibody (BE-Ab2) were markedly elevated in succeeded group after PLII. In contrast, although a significant increase could be observed in the failed group after treatment, the elevation of BE-Ab2 was much lower than that in successful group. PLII therapy significantly up-regulated the percentage of peripheral CD4(+)CD25(+)CD127(-) regulatory T cells (Tregs) in successfully pregnant women; however, there was no significant change of Tregs in pregnancy loss cases although receiving PLII therapy. These results suggested a positive correlation between higher frequency of Tregs and rate of successful pregnancies. The sensitivity and specificity of combination of Tregs with MLC-BE and BE-Ab2 were 81.8% and 81.3%, respectively. Therefore, the percentage of Tregs in peripheral blood may hopefully serve as a potential biomarker for monitoring the efficacy of therapy in URSA patients. Combination of Tregs with MLC-BE and BE-Ab2 may expect to better evaluate the efficacy of PLII in URSA patients.