Identification of End-Binding 1 Protein as Novel α-4 Giardin-Binding Partners in Giardia lamblia Trophozoites.

BACKGROUND: Giardia lamblia (syn. G. intestinalis, G. duodenalis) is a primitive opportunistic protozoon, and one of the earliest differentiated eukaryotes. Despite its primitive nature, G. lamblia has a sophisticated cytoskeleton system, which is closely related to its proliferation and pathogenicity. Meanwhile, α giardin is a G. lamblia-specific cytoskeleton protein, which belongs to the annexin superfamily. Interestingly, G. lamblia has 21 annexin-like α giardins, i.e., more than higher eukaryotes. The functional differences among α giardin members are not fully understood. METHODS: We took α-4 giardin, a member of α giardin family, as a research object. A morpholino-mediated knockdown experiment was performed to identify the effect of α-4 giardin on G. lamblia trophozoites biological traits. A yeast two-hybrid cDNA library of G. lamblia strain C2 trophozoites was screened for interaction partners of α-4 giardin. Co-immunoprecipitation and fluorescent colocalization confirmed the relationship between G. lamblia EB1 (gEB1) and α-4 giardin. RESULTS: α-4 Giardin could inhibit the proliferation and adhesion of G. lamblia trophozoites. In addition, it interacted with G. lamblia EB1 (gEB1). CONCLUSIONS: α-4 Giardin was involved in proliferation and adhesion in G. lamblia trophozoites, and EB1, a crucial roles in mitosis, was an interacting partner of α-4 giardin.

Correlation of Serum IL-17, VEGF, and Lactate Dehydrogenase (LDH) Levels with Prognosis of Gastric Cancer.

Objective: To investigate the correlation of serum interleukin-17 (IL-17), vascular endothelial growth factor (VEGF) and lactate dehydrogenase (LDH) levels with the prognosis of gastric cancer patients. Methods: From December 2018 to December 2020, 45 patients with gastric cancer treated in our hospital and 50 healthy individuals were assessed for eligibility and recruited. The eligible patients were assigned to an observation group, and the healthy subjects were assigned to a control group. Serum IL-17, LDH, and VEGF levels of the eligible participants were determined by the enzyme-linked immunosorbent assay (ELISA) and biochemical testing. The association of serum IL-7, LDH, and VEGF levels with their pathological characteristics was examined in the observation group. The correlation between serum IL-17 and VEGF was analyzed using the Pearson method, and regression models were established using COX proportional risk to explore the independent risk factors for gastric cancer. Results: Gastric cancer was associated with higher levels of IL-17, LDH, and VEGF versus a healthy status (P < 0.05). There was no significant difference in serum IL-17, LDH, and VEGF levels between the two groups of patients with different clinical characteristics (P > 0.05). Higher tumor TNM stages resulted in significantly higher levels of IL-17, LDH, and VEGF (P < 0.05). Serum IL-17 level was positively correlated with VEGF level (P < 0.05). Cox regression multifactorial analysis showed that serum IL-17, LDH, VEGF, and tumor TNM stages could be independent high-risk influencing factors for gastric cancer (P < 0.05). Serum IL-17 was positively correlated with VEGF levels in patients with gastric cancer. Conclusion: Serum IL-17, LDH, and VEGF levels in gastric cancer patients are closely correlated with the TNM stage and patients' prognosis, both of which show great potential as effective indicators for evaluating the prognosis of gastric cancer.

Establishment of a Lung Cancer Discriminative Model Based on an Optimized Support Vector Machine Algorithm and Study of Key Targets of Wogonin in Lung Cancer.

An optimized support vector machine model was used to construct a lung cancer diagnosis model based on serological indicators, and a molecular regulation model of Wogonin, a component of Scutellaria baicalensis, was established. Serological indexes of patients were collected, the grid search method was used to identify the optimal penalty coefficient C and parameter g of the support vector machine model, and the benign and malignant auxiliary diagnosis model of isolated pulmonary nodules based on serological indicators was established. The regulatory network and key targets of Wogonin in lung cancer were analyzed by network pharmacology, and key targets were detected by western blot. The relationship between serological susceptibility genes and key targets of Wogonin was established, and the signaling pathway of Wogonin regulating lung cancer was constructed. After support vector machine parameter optimization (C = 90.597, g = 32), the accuracy of the model was 90.8333%, with nine false positives and two false negative cases. Ontology functional analysis of 67 common genes between Wogonin targets and lung cancer-related genes showed that the targets were associated with biological processes involved in peptidye-serine modification and regulation of protein kinase B signaling; cell components in the membrane raft and chromosomal region; and molecular function in protein serine/threonine kinase activity and heme binding. Kyoto Encyclopedia of Genes and Genomes analysis showed that the regulation pathways involved the PI3K-Akt signaling pathway, ERBB signaling pathway, and EGFR tyrosine kinase inhibitor resistance. In vitro analyses using lung cancer cells showed that Wogonin led to significantly increased levels of cleaved caspase-3 and Bad and significantly decreased Bcl-2 expression in a concentration-dependent manner. ErbB4 expression also significantly decreased in lung cancer cells after treatment with Wogonin. A regulatory network of Wogonin regulating lung cancer cell apoptosis was constructed, including the participation of serological susceptibility genes. There is a certain regulatory effect between the serological indexes that can be used in the diagnosis of lung cancer and the key targets of Chinese herbal medicine treatment of lung cancer, which provides a new idea for the diagnosis, treatment and prognosis of clinical lung cancer.

[Establishment of dual reverse phase chromatography combined parallel reaction monitoring for targeted quantitative proteomics].

Steady improvement in mass spectrometers technology has transformed the targeted proteome analysis into a new stage. Parallel reaction monitoring (PRM) technology has evolved from the basic multiple reaction monitoring (MRM) targeted proteomics methods in recent years. PRM performs with a higher sensitivity, throughput and reproducibility in targeted quantification, however its limitations in effectiveness and accurate quantification of samples with higher complexity still remain unsolved. In this study through improving the chromatographic conditions of PRM we established a simple and robust platform for targeted proteomic quantification. The newly established PRM system is equipped with columns with increased inner diameter (150 µm) and decreased total length (8 cm); faster liquid phase elution rate (800 nL/min) and shortened elution gradient (35 min). These modifications enable PRM platform to combine with dual reverse phase chromatography, to quantify up to 400 low abundance peptides in human 293T cells whole cell extract. Our findings would benefit the promotion of PRM technology, especially providing a technical option for accurate quantification of low abundance proteins.

The mediating role of emotional intelligence between negative life events and psychological distress among nursing students: A cross-sectional study.

BACKGROUND: Previous studies have highlighted that negative life events and emotional intelligence are significant predictors of mental health. However, whether emotional intelligence mediates the relationship between negative life events and psychological distress among nursing students have not been given adequate attention. OBJECTIVES: To explore the relationship among negative life events, emotional intelligence and psychological distress and to examine the mediating role of emotional intelligence in psychological distress among Chinese nursing students. DESIGN: A cross-sectional survey using convenience sampling. SETTINGS AND PARTICIPANTS: A total of 467 nursing students who were enrolled in a university in mainland of China. METHODS: A structured questionnaire was administered from September-November in 2013 to participants who consented to participate in the study. Independent variables were personal variables, emotional intelligence and negative life events. Outcome variable was psychological health. The means and standard deviations were computed. Student's t-test and one-way analysis of variance (ANOVA) were performed, to test the differences among the demographic characteristics on the psychological distress scores. Pearson correlation analyses and hierarchical regression analyses were performed. RESULTS: Negative life events were positively associated with psychological distress. Emotional intelligence was negatively associated with psychological distress and negative life events. Emotional intelligence mediated the relationship between negative life events and psychological distress. CONCLUSIONS: The findings support the theory of Salovey and his colleagues, and provide evidence for emotional intelligence as a factor that buffers effects of negative life events on psychological distress.

Co-circulation of Hantaan, Kenkeme, and Khabarovsk Hantaviruses in Bolshoy Ussuriysky Island, China.

Hemorrhagic fever with renal syndrome (HFRS) was first recognized in far eastern Asia in the 1930s, and has been highly prevalent in this region ever since. To reveal the molecular epidemiology of hantaviruses in this region, a total of 374 small mammals (eight species of rodents and one species of shrew) were captured in the Chinese part of the Bolshoy Ussuriysky Island (Heilongjiang Province). Hantavirus sequences were recovered from three striped field mice (Apodemus agrarius), 11 Maximowicz's voles (Microtus maximowiczii), and one flat-skulled shrew (Sorex roboratus). Genetic and phylogenetic analysis revealed the presence of three viruses: Hantaan virus (HTNV), Khabarovsk virus (KHAV), and Kenkeme virus (KKMV). HTNV sequences recovered from A. agrarius were closely related to those identified in Apodemus mice from the surrounding areas, while a new lineage of KHAV was present in M. maximowiczii. Additionally, while the viral sequences recovered from one flat-skulled shrew were most closely related to KKMV, their divergence to the prototype strain suggests that they represent a new viral subtype. Overall, these results suggest that Bolshoy Ussuriysky Island harbors considerable hantavirus diversity.

[Changes of IFN-gamma, IL-4 and T cells in Schistosoma japonicum-infected mice after praziquantel treatment].

OBJECTIVE: To investigate the serum levels of IFN-gamma and IL-4, and the dynamic changes of IFN-gamma-specific and IL-4-specific lymphocytes in mice with Schistosoma japonicum infection after treatment by praziquantel. METHODS: Ninety BALB/c mice were randomly divided into three groups (n = 30) named as infection group, treatment group and control group. The mice in treatment group and infection group were infected with (25 +/- 2) S. japonicum cercariae through the abdominal skin. At 6 weeks post-infection, the mice in treatment group were administered orally with praziquantel [300 mg/(kg x d)] for 3 d. At 4, 6, 8 and 12 weeks post-treatment, the mice were weighed, and serum samples were collected. Serum levels of IFN-gamma and IL-4 were measured by ELISA. At the same time, the spleens were aseptically removed to prepare cell suspension, and the counts of IFN-gamma and IL-4 specific lymphocytes were examined by ELISPOT after stimulation of Schistosoma japonicum soluble egg antigen (SEA). RESULTS: From 4 to 12 weeks after praziquantel treatment, the body weight of mice in treatment group were significantly heavier than that of infection group (P < 0.05), but no significant difference was found between treatment group and control group (P < 0.05). At 4 weeks posttreatment, there was no significant difference in serum levels of IFN-gamma and IL-4 between treatment group and infection group (P > 0.05). At 6, 8, and 12 weeks after treatment, the serum levels of IFN-gamma (0.038 +/- 0.013, 0.028 +/- 0.001, and 0.027 +/- 0.007) and IL-4(0.051 +/- 0.020, 0.045 +/- 0.019, and 0.043 +/- 0.016) in treatment group were significantly lower than that of infection group (IFN-gamma: 0.057 +/- 0.004, 0.060 +/- 0.023, and 0.052 +/- 0.017; IL-4: 0.150 +/- 0.014, 0.148 +/- 0.014, and 0.123 +/- 0.017) (P < 0.05). Serum IFN-gamma and IL-4 levels in treatment group and infection group were significantly higher than that of control group (P < 0.05). ELISPOT results showed that at 4, 6 weeks post-treatment, there was no significant difference in the number of IFN-gamma-specific lymphocytes between treatment group and infection group (P > 0.05). While at 8 and 12 weeks after treatment, the IFN-gamma-specific lymphocytes in treatment group (39.9 +/- 22.8 and 38.5 +/- 6.2) were significantly less than that of infection group (141.9 +/- 39.3 and 106.8 +/- 28.6) (P < 0.05). At 4-week post-treatment, the IL-4-specific lymphocytes in treatment group were much more than that of infection group (175.6 +/- 62.3) (P < 0.05), and then began to decline. At 8 and 12 weeks after treatment, the IL-4-specific lymphocytes (111.3 +/- 14.3 and 113.0 +/- 44.2) in treatment group were significantly less than that of infection group (220.3 +/- 107.1 and 208.1 +/- 17.2) (P < 0.05). The IFN-gamma-specific and IL-4-specific lymphocytes in treatment group and infection group were significantly more than that of control group (P < 0.05). CONCLUSION: After praziquantel treatment, the serum levels of IFN-gamma and IL-4 in mice with S. japonicum infection decrease, and the number of IFN-gamma and IL-4 specific lymphocytes reduces.

Construction and characterization of a full-lengh cDNA library from non-fresh Giardia lamblia.

OBJECTIVE: To construct rapidly a full-length cDNA library from nanogram amounts total RNA of Giardia lamblia (G. lamblia) trophozoites stocked in RNA stabilization reagent. METHODS: Total RNA of Giardia was extracted using Trizol reagent. A full-length cDNA library of G. lamblia trophozoites was constructed by a long-distance PCR (LD-PCR) method. The recombinant rate and the coverage rate of full-length clones of the library were evaluated. The inserted fragments were identified and sequenced by PCR amplification. RESULTS: The titer of cDNA library was 3.85 × 10(7) pfu/mL. The length of inserted fragments ranged from 0.4 to 2.5 kb, and the recombination efficiency accounted for 100% (20/20). The coverage rate of full-length clones is high (17/20). CONCLUSIONS: The RNA stabilization reagent may be used to fix the cells and prevent the RNA in cells even though delivered under normal atmospheric temperature. The long-distance PCR can be used to construct a full-length cDNA library rapidly and it needs less RNA than the traditional method from mRNA.

Toll-like receptor 5 agonism protects mice from radiation pneumonitis and pulmonary fibrosis.

Radiation pneumonitis and pulmonary fibrosis are the main complications with radiotherapy for thoracic neoplasms, directly limiting the efficient dose in clinical application and currently there are few medicines that effectively function as radioprotectants. However, a TLR5 agonist, CBLB502, was confirmed to have protective efficacy against hematopoietic and gastrointestinal radiation syndromes in mice and primates. This study points to a new direction for protection against thoracic radiation-induced pulmonary syndromes and skin injury by CBLB502. We utilized the TUNEL assay, pathological analysis and immunohistochemistry to obtain evidence that CBLB502 could alleviate the occurrence of radiation pneumonitis and pulmonary fibrosis as well as radiation- induced skin injury. It may thus play a promising role in facilitating clinical radiotherapy of thoracic neoplasms.

[Construction of GCV-specific hammerhead ribozyme recombinant vector of alpha-8 giardin in Giardia lamblia].

OBJECTIVE: To construct a GCV-ribozyme recombinant vectors of alpha-8 giardin in Giardia lamblia. METHODS: The secondary structure of alpha-8 giardin mRNA (GenBank Accession No. AY781323) was analyzed with the RNA draw software. According to the proportion of G:C and principles of designing hammerhead ribozyme, suitable ribozyme cleavage points were chosen. A specific antisense-hammerhead ribozyme (H8) was designed and synthesized. The ribozyme was cloned into Giardia canis virus (GCV) vector to construct a recombinant viral vector-pGCV634/H8/1423. The vector was linearized and transcript into the trophozoites of G. lamblia by electroporation method. The alpha-8 giardin mRNA level of the transfectants and normal trophozoites were analyzed 24 h after electroporation by RT-PCR. RESULTS: The recombinant vector of GCV-specific hammerhead ribozyme of alpha-8 giardin in Giardia lamblia (pGCV634/H8/ 1423) was constructed. RT-PCR assays showed the ribozyme (H8) mRNA can be detected 24h after transfection and alpha-8 giardin mRNA was cleaved effectively by ribozyme (H8) intracellularly. CONCLUSION: pGCV634/H8/1423 can transfect Giardia trophozoites and cleave mRNA of alpha-8 giardin intracellularly.

[Facial Demodex infection among college students in Tangshan].

A survey indicated that the prevalence of Demodex infection among 512 college students in Tangshan was 36.3% (186/512), that of males and females was 39.3% (81/206) and 34.3% (105/306) respectively (P>0.05). The infection of Demodex folliculorum accounted for 82.3% (153/186), followed by D. brevis (7.5%, 14/186) and mixed infection (10.2%, 19/186). The prevalence was 47.0% (93/198) in subjects with oily skin, 26.6% (37/139) in those with dry skin, and 33.9% (56/165) in mixed-type skin (P<0.05). Subjects with facial diseases (62.0%, 75/121), such as rosacea and acne, were more likely to be infected with Demodex than those with healthy skin (27.6%, 80/290) (P<0.05). Prevalence in those lived in humid environment (67.9%, 95/140) was higher than those lived in the desiccating environment (24.5%, 91/372) (P<0.05).

[Entamoeba gingivalis infection among college students in Tangshan].

OBJECTIVE: To investigate the prevalence of Entamoeba gingivalis infection in college students in Tangshan, and analyze the relationship between the infection and human behaviors. METHODS: 551 students of grades 1-3 from six colleges in Tangshan received questionnairing, which covered the oral health state, teeth-brushing, xylitol gum-chewing, diet fondness, and smoking. Specimens were taken from the tooth surface of the lesion or fouling materials by using disinfected toothpicks and the smears were observed microscopically to examine Entamoeba gingivalis infection. RESULTS: The prevalence of Entamoeba gingivalis infection was 28.3% (156/551), 30.4% (55/181) and 24.6% in males and females (91/370) respectively (chi2=2.09, P>0.05). The prevalence in students with or without oral disorders was 41.2% (84/204) and 20.8% (72/347) respectively, with a significant statistical difference (chi2=26.41, P<0.01); it was 22.5% (53/236) and 32.7% (103/315) among students who cleaned their teeth regularly or irregularly (chi2=6.97, P<0.01); it was 18.3% (17/93) and 30.4% (139/458) among those usually with or without chewing xylitol gum (chi2=5.55, P<0.05). CONCLUSION: Entamoeba gingivalis infection is common in the college students in Tangshan and it has a close relation to the oral hygiene habits and the presence of oral disorders.

[Human parasitology teaching in the 21st century].

Multimedia techniques were applied in parasitological teaching and experimental practices. In order to strengthen the practical ability of the undergraduate students, reform was conducted including a prioritization of the teaching content, use of series micro-slides and video show, etc.

[The injury of metronidazole on morphology of Giardia lamblia in vitro].

Trophozoites of Giardia lamblia were axenically cultivated with modified TYI-S-33 medium contained 500 microg/ml metronidazole (12h LC50). The morphology of drug-treated trophozoites was observed with light and electron micro-scopes at 2, 4, 8, 12 h respectively. The light microscopy revealed that the trophozoites treated with MTZ showed swollen, detached from the wall of the culture tube, and were with vacuoles in the cytoplasm. Movement of the flagella become slowly or stopped. Electronic microscopy showed that the trophozoites were swollen and deformed; lots of vacuoles were seen in the cytoplasm; the contents of cytoplasm were depleted and the nuclei deformed. This study indicated that MTZ has injured the morphology of G. lamblia.

[Effect of dihydroartemisinin on ultrastructure of Giardia lamblia in vitro].

OBJECTIVE: To study the in vitro effect of dihydroartemisinin (DHA) on Giardia lamblia. METHODS: Trophozoites of G. lamblia were cultivated with modified TYI-S-33 medium that contains dihydroartemisinin (DHA). The trophozoites were morphologically observed respectively with light and electron microscopes after treated with the drug. RESULTS: The mortality increased with the prolongation of the time of the drug action and the increase of drug concentration (P<0.01). While at the same concentration of 100 microg/ml, the mortality increased from 46.6% for 12 h to 100% for 24 h (P<0.05). For 12 h, the mortality of G. lamblia was from 46.6% at concentration of 100 microg/ml to 100% at 200microg/ml. Under optical microscope, deformation and swelling of the parasites were observed when treated with DHA for 12, 24 and 48 h. Movement of the flagella became slow or stopped. Under electron microscope, the trophozoites were swollen and deformed, vacuoles were seen in the cytoplasm, and the cell membrane ruptured and fell off. The cytoplasm protrusions appeared on the surface of the plasma membrane. The adhesive disc changed into large bubbles and the perinuclear space became wider and the deformed nucleus was seen. CONCLUSION: DHA shows a strong impairment on the plasma membrane and cytoskeleton of Giardia lamblia.