[Construction of RNA interfering lentiviral vector of occludin and its role in tight junctions].

Objective: To investigate the role of occludin in tight junction (TJ) in vitro.

Methods: We constructed RNA interfering lentiviral vectors and transfected them into TM4 cells. Then we detected their inhibitory effect on occuldin by RT-PCR and Western blot and analyzed the role of occuldin in TJ using an in vitro TJ cell model.

[SP1 identified as a transcription factor of miR-122-5p].

Objective: To study the transcription factors of the spermatogenesis-related promoter mir-122-5p.

Methods: SP1 and GATA4 were predicted as the possible transcription factors of the mir-122-5p promoter by bioinformatics analysis, followed by construction of the double luciferase pGL3-mir-122-5p promoter vector, pcDNA3.1 (+) -SP1 expression vector and pcDNA3.

miR-122-5p regulates the tight junction of the blood-testis barrier of mice via occludin : miR-122-5p can regulate the tight junction.

Background: Occludin protein is the primary assembling protein of TJs and the structural basis for tight junction formation between Sertoli cells in the spermatogenic epithelium. The expression of miR-122-5p and occludin are negatively correlated. In order to investigate the regulation mechanism of miR-122-5p on occludin and TJ, the present study isolated primary Sertoli cells from C57BL/6 mice, identified a transcription factor of miR-122-5p in testicle, studied the modulating loci of miR-122-5p on occludin using a dual-luciferase reporter assay, analyzed the regulate of miR-122-5p on the expression of occludin with real-time RT-PCR and Western blot, and studied the effect of miR-122-5p on the tight junction using a Millicell Electrical Resistance System.

A 22-amino-acid peptide regulates tight junctions through occludin and cell apoptosis.

Occludin is a structural protein of tight junctions (TJ) in the blood-testis barrier (BTB). A 22-amino-acid peptide (22AA) in the second extracellular loop can reversibly regulate TJ, but its regulatory mechanism is unknown. In this study, a 22AA-induced TJ destruction animal model was constructed to investigate the effect of 22AA on Sertoli cells (SCs) and spermatid counts and cell apoptosis at different time points using a multiplex immunofluorescence technique.

Correlation Analysis of miR-122-5p and Occludin with Sperm Density in Oligospermia Patients' Sperm.

Background: To analyze the correlation of miR-122-5p and occludin with sperm density in oligospermia patients' sperm.

Methods: The expression of miR-122-5p and its target protein occludin in the sperm and exfoliated cells of semen were studied using real-time reverse transcription-polymerase chain reaction and Western blot analysis.

Results: The expression level of miR-122-5p in the sperm and exfoliated cells of patients with oligospermia semen was 0.

[Cross immunogenicity between Ia-associated invariant chains of chicken (Gallus domestiaus) and Muscovy duck (Cairina moschata)].

Objective: To explore the cross immunogenicity between Ia-associated invariant chain (Ii) of chicken (CIi) and Ii of Muscovy duck (MDIi).

Methods: The immunoreactivity between mouse anti-CIi serum and the MDIi expressed by prokaryotic cell, and the immunoreactivity between the mouse anti-MDIi serum and the CIi expressed by prokaryotic cell were detected through indirect ELISA. To detect the immunoreactivity between the mouse anti-CIi serum and the MDIi of the spleen tissues of Muscovy duck, and the immunoreactivity between the mouse anti-MDIi serum and the CIi of the liver tissues of chicken (tissular CIi), the fluorescence immunohistochemistry assay was performed using DyLight 488 affiniPure goat anti-mouse IgG as the second antibody.

[Preparation and characterization of polyclonal antibody against Ia-associated invariant chain of Muscovy Duck (Cairina moschata)].

Objective: To prepare polyclonal antibody against invariant chain of Muscovy duck (Cairina moschata) (MDIi) and identify its reaction with MDIi extracted from tissues of Muscovy duck.

Methods: MDIi was amplified by PCR and used to construct the prokaryotic expression vector of pET-32a/MDIi by linking with the plasmid of pET-32a. Then pET-32a/MDIi was transformed into E.

Single-chain antibody against human lipocalin-type prostaglandin D synthase: construction, expression, purification, and activity assay.

An active form of single-chain antibody (ScFv) from murine monoclonal antibody 4A7, which is specific for lipocalin-type prostaglandin D synthase (L-PGDS), was produced in Escherichia coli. The complementary DNA fragments encoding the variable regions of heavy chain (VH) and light chain (VL), which amplified from hybridoma 4A7 producing a monoclonal antibody (IgG1) against L-PGDS, were connected by a (Gly4Ser)3 linker using an assembly polymerase chain reaction. The resultant ScFv were cloned into the vector pGEM and expressed in E.

Relationship between contents of lipocalin-type prostaglandin D synthase on the surface of infertility sperm and in seminal plasma.

Lipocalin-type prostaglandin D synthase (L-PGDS) is localized in Leydig cells, sperm, and epithelial cells of the epididymis. The present study was to determine the correlation between content of this enzyme in seminal plasma and on the surface of sperm. We analyzed 90 semen samples.

Different expression of lipocalin-type prostaglandin D synthase in rat epididymidis.

This study was designed to explore the different expression of L-PGDS (lipocalin-type prostaglandin D synthase) in rat epididymidis and to gain further insight into the potential function of L-PGDS in male reproduction. The expression of L-PGDS in rat epididymidis was assessed using real-time quantitative PCR and immunoblotting. The distribution of L-PGDS in rat epididymidis was explored by immunohistochemical methods.