scholarly journals Characterization and Localization of Calb2 in Both the Testis and Ovary of the Japanese Flounder (Paralichthys olivaceus)

Animals ◽  
2020 ◽  
Vol 10 (9) ◽  
pp. 1503
Author(s):  
Yuting Xiang ◽  
Yahui Wu ◽  
Haoran Zhang ◽  
Jikui Wu ◽  
Junling Zhang
Keyword(s):  
Real Time ◽  
Western Blotting ◽  
Real Time Pcr ◽  
Paralichthys Olivaceus ◽  
Neighboring Gene ◽  
Syntenic Relationship ◽  
Reading Frame ◽  
Cdna Sequences ◽  
Its Gene ◽  
And Function

Although its function in mammalian gonads has been gradually recognized, the expression and function of calretinin (CALB2)—a Ca2+-binding protein—in the testis and ovary of fish are still unclear. Here, we identified the cDNA sequences of calb2 in Paralichthys olivaceus (P. olivaceus); analyzed its gene structure and phylogenetic and syntenic relationship by bioinformatics; and investigated its tissue distribution and localization in the gonads by real-time PCR, western blotting, and immunohistochemistry. The P. olivaceuscalb2 gene has 11 exons and 10 introns, and the full-length cDNA is 1457 bp, including an open reading frame (ORF) of 816 bp encoding 271 amino acids. The CALB2 of P. olivaceus has a higher homology with Lates calcarifer (99%) compared with other species. The conserved synteny of calb2 neighboring gene loci was also detected in fish. Real-time PCR showed that the expression of calb2 mRNA is abundant not only in the brain, but also in the gonads, and exhibits a higher expression in the testis than in the ovary. Western blotting indicated that the CALB2 protein has a higher expression in the testis compared with the ovary. Immunohistochemistry demonstrated that the CALB2 protein appears in Leydig cells and the ovarian germ epithelium. These results reveal that calb2 plays an important role in the gonads of P. olivaceus.

Increased Expression of TXNIP Facilitates Oxidative Stress in Nasal Epithelial Cells of Patients With Chronic Rhinosinusitis With Nasal Polyps

2020 ◽  
pp. 194589242098241
Author(s):  
Hai Lin ◽  
Guangyi Ba ◽  
Ru Tang ◽  
Mingxian Li ◽  
Zhipeng Li ◽  
...  
Keyword(s):  
Oxidative Stress ◽  
Epithelial Cells ◽  
Real Time ◽  
Chronic Rhinosinusitis ◽  
Western Blotting ◽  
Real Time Pcr ◽  
Nasal Polyps ◽  
Sod Activity ◽  
Nasal Tissue ◽  
Sod Assay

Background Oxidative stress plays crucial roles in the pathogenesis of chronic rhinosinusitis with nasal polyps (CRSwNP). Thioredoxin-interacting protein (TXNIP) is essential in the process of triggering oxidative stress. However, its role and mechanism in CRSwNP remain unclear. The present study sought to explore the role and mechanism of TXNIP in the pathogenesis of CRSwNP. Methods Western blotting, real-time PCR and immunohistochemistry (IHC) were employed to assess TXNIP, thioredoxin (TRX) expression in nasal tissue samples from patients with CRSwNP and control subjects. MDA level and SOD activity in nasal tissue homogenates were measured using MDA and SOD Assay Kit. To evaluate the role and mechanism of TXNIP in CRSwNP, human nasal epithelial cells (HNECs) were cultured and stimulated using TXNIP siRNA, with or without N-acetylcysteine (NAC, an ROS scavenger). Western blotting, real-time PCR, ROS detecting dye DCFH-DA, MDA and SOD Assay Kit were performed to assess the effects and mechanisms of stimulators on the cells. Results We found significantly increased levels of TXNIP and decreased levels of TRX protein, mRNA, positive cells, increased MDA level and decreased SOD activity in CRSwNP patients compared with control subjects. In vitro study, significantly altered levels of TXNIP, TRX, MDA, SOD and ROS in HNECs were found following treatment of TXNIP siRNA with or without NAC on HNECs. Conclusion TXNIP expression was increased and TRX expression was decreased in CRSwNP at both protein and mRNA levels. MDA levels were increased and SOD activities were decreased in CRSwNP. TXNIP may have negative association with TRX, and then decrease SOD activities and increase MDA levels, resulting in the upregulation of ROS and oxidative stress in HNECs, which may play a pivotal role in the pathogenesis of CRSwNP. Future studies are expected to further explore the role and mechanism of TXNIP in CRSwNP.

scholarly journals Protein kinase Cδ expression in breast cancer as measured by real-time PCR, western blotting and ELISA

2008 ◽  
Vol 99 (10) ◽  
pp. 1644-1650 ◽  
Author(s):  
E McKiernan ◽  
K O'Brien ◽  
N Grebenchtchikov ◽  
A Geurts-Moespot ◽  
A M Sieuwerts ◽  
...  
Keyword(s):  
Breast Cancer ◽  
Protein Kinase ◽  
Real Time ◽  
Western Blotting ◽  
Real Time Pcr ◽  
Protein Kinase Cδ

scholarly journals Detection of Vibrio parahaemolyticus in Shellfish by Use of Multiplexed Real-Time PCR with TaqMan Fluorescent Probes

2006 ◽  
Vol 72 (3) ◽  
pp. 2031-2042 ◽  
Author(s):  
Linda N. Ward ◽  
Asim K. Bej
Keyword(s):  
Real Time ◽  
Real Time Pcr ◽  
Vibrio Parahaemolyticus ◽  
Detection Methods ◽  
Tissue Homogenate ◽  
Oyster Tissue ◽  
Reading Frame ◽  
Initial Inoculum ◽  
Hemolysin Gene ◽  
Pure Cultures

ABSTRACT We developed a multiplexed real-time PCR assay using four sets of gene-specific oligonucleotide primers and four TaqMan probes labeled with four different fluorophores in a single reaction for detection of total and pathogenic Vibrio parahaemolyticus, including the pandemic O3:K6 serotype in oysters. V. parahaemolyticus has been associated with outbreaks of food-borne gastroenteritis caused by the consumption of raw or undercooked seafood and therefore is a concern to the seafood industry and consumers. We selected specific primers and probes targeting the thermostable direct hemolysin gene (tdh) and tdh-related hemolysin gene (trh) that have been reported to be associated with pathogenesis in this organism. In addition, we targeted open reading frame 8 of phage f237 (ORF8), which is associated with a newly emerged virulent pandemic serotype of V. parahameolyticus O3:K6. Total V. parahaemolyticus was targeted using the thermolabile hemolysin gene (tlh). The sensitivity of the combined four-locus multiplexed TaqMan PCR was found to be 200 pg of purified genomic DNA and 104 CFU per ml for pure cultures. Detection of an initial inoculum of 1 CFU V. parahaemolyticus per g of oyster tissue homogenate was possible after overnight enrichment, which resulted in a concentration of 3.3 × 109 CFU per ml. Use of this method with natural oysters resulted in 17/33 samples that were positive for tlh and 4/33 samples that were positive for tdh. This assay specifically and sensitively detected total and pathogenic V. parahaemolyticus and is expected to provide a rapid and reliable alternative to conventional detection methods by reducing the analysis time and obviating the need for multiple assays.

Increased relaxation of immature airways to β2-adrenoceptor agonists is related to attenuated expression of postjunctional smooth muscle muscarinic M2 receptors

2005 ◽  
Vol 98 (4) ◽  
pp. 1526-1533 ◽  
Author(s):  
Michael Fayon ◽  
Eric Dumas De La Roque ◽  
Patrick Berger ◽  
Hugues Begueret ◽  
Olga Ousova ◽  
...  
Keyword(s):  
Smooth Muscle ◽  
Real Time ◽  
Airway Smooth Muscle ◽  
Western Blotting ◽  
Real Time Pcr ◽  
Molecular Techniques ◽  
Receptor Expression ◽  
Smooth Muscle Relaxation ◽  
Fetal Life ◽  
Mrna Levels

Spontaneous or agonist-induced contraction of airway smooth muscle can be observed very early in fetal life, thus explaining the possible occurrence of bronchospasm in very low birth weight infants within the first days of life. In an attempt to better manage such bronchospasms, the aim of the present study was to investigate the age-specific modifications in airway smooth muscle relaxation to β2-agonists and muscarinic antagonists using a combination of functional and molecular techniques. In the rat, isometric relaxation to the β2-agonist salbutamol was examined in tracheae; we also examined muscarinic receptor expression (M2R and M3R mRNA levels) in airway smooth muscle by immunochemistry, Western blotting, and real-time PCR. Compared with adults, salbutamol-induced relaxation was twofold greater in immature rat isolated tracheae preconstricted by carbachol. This effect was associated with a lower expression of M2R in the smooth muscle of immature animals (sixfold and almost twofold as assessed by immunochemistry and Western blotting, respectively). Real-time PCR data indicate that changes in M2R expression according to age occurred at a posttranscriptional level. In adult airways, there was a significantly greater functional efficacy of M2R blockade by methoctramine compared with that shown in immature rats. Because of the limited availability of human neonate lung tissue, only the molecular part of the study was performed, and we observed a qualitatively similar effect, i.e., a lower M2R expression in the neonatal airway smooth muscle, although this was quantitatively smaller. We conclude that β2-agonist-induced relaxation is enhanced in immature compared with adult airways as a result of greater postjunctional M2R expression in adult airway smooth muscle. This finding may be of importance in the clinical management of bronchoconstriction in neonates.

Sepsis stimulates calpain activity in skeletal muscle by decreasing calpastatin activity but does not activate caspase-3

2005 ◽  
Vol 288 (3) ◽  
pp. R580-R590 ◽  
Author(s):  
Wei Wei ◽  
Moin U. Fareed ◽  
Amy Evenson ◽  
Michael J. Menconi ◽  
Hongmei Yang ◽  
...  
Keyword(s):  
Skeletal Muscle ◽  
Real Time ◽  
Western Blotting ◽  
Real Time Pcr ◽  
Caspase 3 ◽  
Calpain Inhibitor ◽  
Mrna Levels ◽  
Protein Breakdown ◽  
Calpain Activity ◽  
Hydrolysis Of

We examined the influence of sepsis on the expression and activity of the calpain and caspase systems in skeletal muscle. Sepsis was induced in rats by cecal ligation and puncture (CLP). Control rats were sham operated. Calpain activity was determined by measuring the calcium-dependent hydrolysis of casein and by casein zymography. The activity of the endogenous calpain inhibitor calpastatin was measured by determining the inhibitory effect on calpain activity in muscle extracts. Protein levels of μ- and m-calpain and calpastatin were determined by Western blotting, and calpastatin mRNA was measured by real-time PCR. Caspase-3 activity was determined by measuring the hydrolysis of the fluorogenic caspase-3 substrate Ac-DEVD-AMC and by determining protein and mRNA expression for caspase-3 by Western blotting and real-time PCR, respectively. In addition, the role of calpains and caspase-3 in sepsis-induced muscle protein breakdown was determined by measuring protein breakdown rates in the presence of specific inhibitors. Sepsis resulted in increased muscle calpain activity caused by reduced calpastatin activity. In contrast, caspase-3 activity, mRNA levels, and activated caspase-3 29-kDa fragment were not altered in muscle from septic rats. Sepsis-induced muscle proteolysis was blocked by the calpain inhibitor calpeptin but was not influenced by the caspase-3 inhibitor Ac-DEVD-CHO. The results suggest that sepsis-induced muscle wasting is associated with increased calpain activity, secondary to reduced calpastatin activity, and that caspase-3 activity is not involved in the catabolic response to sepsis.

scholarly journals Molecular Cloning and Sequence Analysis of the cDNAs Encoding Toxin-Like Peptides from the Venom Glands of Tarantula Grammostola rosea

2012 ◽  
Vol 2012 ◽  
pp. 1-10 ◽  
Author(s):  
Tadashi Kimura ◽  
Seigo Ono ◽  
Tai Kubo
Keyword(s):  
Sequence Analysis ◽  
Molecular Cloning ◽  
Real Time ◽  
Cdna Library ◽  
Real Time Pcr ◽  
Bioactive Peptides ◽  
Venom Gland ◽  
Unique Peptide ◽  
Cdna Sequences ◽  
Venom Glands

Tarantula venom glands produce a large variety of bioactive peptides. Here we present the identification of venom components obtained by sequencing clones isolated from a cDNA library prepared from the venom glands of the Chilean common tarantula, Grammostola rosea. The cDNA sequences of about 1500 clones out of 4000 clones were analyzed after selection using several criteria. Forty-eight novel toxin-like peptides (GTx1 to GTx7, and GTx-TCTP and GTx-CRISP) were predicted from the nucleotide sequences. Among these peptides, twenty-four toxins are ICK motif peptides, eleven peptides are MIT1-like peptides, and seven are ESTX-like peptides. Peptides similar to JZTX-64, aptotoxin, CRISP, or TCTP are also obtained. GTx3 series possess a cysteine framework that is conserved among vertebrate MIT1, Bv8, prokineticins, and invertebrate astakines. GTx-CRISP is the first CRISP-like protein identified from the arthropod venom. Real-time PCR revealed that the transcripts for TCTP-like peptide are expressed in both the pereopodal muscle and the venom gland. Furthermore, a unique peptide GTx7-1, whose signal and prepro sequences are essentially identical to those of HaTx1, was obtained.

scholarly journals Μοριακή παθολογία νεφρικής ίνωσης

2012 ◽  
Author(s):  
Φανή Καραγιάννη
Keyword(s):  
Real Time ◽  
Western Blotting ◽  
Real Time Pcr ◽  
Rt Pcr ◽  
Galectin 3

Είναι ευρέως γνωστό ότι η Χρόνια Νεφρική Νόσο (ΧΝΝ) είναι μία κατάσταση που εντοπίζεται σε μεγάλη συχνότητα στους ενήλικες (de Jong et al, 2008). Αν και η ΧΝΝ είναι το τελικό αποτέλεσμα αρκετών νεφρικών και συστημικών ασθενειών, το πιο κοινό χαρακτηριστικό της, ανεξάρτητα από την αιτιολογία, είναι η ανάπτυξη νεφρικής ίνωσης. Οι μοριακοί μηχανισμοί και τα μακρομόρια που εμπλέκονται στην ανάπτυξη της νεφρικής ίνωσης δεν έχουν μελετηθεί πλήρως. Για το λόγο αυτό, είναι πολύ σημαντικό να κατανοηθούν οι μοριακοί μηχανισμοί που συμβαίνουν κατά τη διάρκεια της ινωτικής διαδικασίας και να βρεθούν πρώιμοι μάρτυρες για την έγκαιρη ανίχνευσή της.Η ίνωση ορίζεται συχνά ως μια διαδικασία επούλωσης τραύματος που έχει διαφύγει από τον έλεγχο του οργανισμού, χαρακτηρίζοντας μεγάλο εύρος ασθενειών σε πολλά οργανικά συστήματα. Κύριο γνώρισμα της ίνωσης αποτελεί η συσσώρευση κυττάρων ινοβλαστών και η ανεξέλεγκτη παραγωγή εξωκυττάριας ουσίας που οδηγεί σε απώλεια της δομής και λειτουργίας του ιστού, και σε πολλές περιπτώσεις οργανική ανεπάρκεια και θάνατο.Με σκοπό την πληρέστερη εξέταση των μορίων που εμπλέκονται στη νεφρική ίνωση, πραγματοποιήσαμε πρωτεωμική ανάλυση του νεφρικού φλοιού στο καθιερωμένο μοντέλο της μονόπλευρης απόφραξης ουρητήρα στον αρουραίο (UUO model). Μεταξύ των πρωτεϊνών που έδειξαν διαφορική έκφραση ανάμεσα στα ζώα μάρτυρες και στα ινωτικά ζώα είναι η τρανσγελίνη. Η τρανσγελίνη, μία πρωτεΐνη του κυτταροσκελετού που φυσιολογικά συναντάται μόνο στα λεία μυϊκά κύτταρα, παρουσίασε σημαντική αύξηση σε όλα τα πειραματικά ζώα που θυσιάστηκαν οχτώ ημέρες μετά την απόφραξη του ουρητήρα.Ο σκοπός της παρούσας μελέτης ήταν, από την μία μεριά, η επιβεβαίωση της υπερέκφρασης της τρανσγελίνης, ο εντοπισμός της στον νεφρικό ιστό, ο χαρακτηρισμός των κυττάρων που την εκφράζουν και είναι υπεύθυνα για την αύξηση της έκφρασής της καθώς επίσης και ο μηχανισμός υπερέκφρασης της τρανσγελίνης στο ινωτικό μοντέλο της μονόπλευρης απόφραξης ουρητήρα σε τρωκτικό, και από την άλλη η μελέτη αρκετών νεφρικών ασθενειών στον άνθρωπο για να διαπιστωθεί η έκφραση της τρανσγελίνης στο νεφρικό ινωτικό ιστό ασθενών.Η υπερέκφραση της τρανσγελίνης μελετήθηκε τόσο σε επίπεδο πρωτεϊνης με Western Blotting, όσο και σε επίπεδο mRNA με RT-PCR. Ο εντοπισμός της τρανσγελίνης στο νεφρικό ιστό μετά την ανάπτυξη της ίνωσης πραγματοποιήθηκε με πειράματα ανοσοϊστοχημείας. Για να καθοριστεί το είδος των κυττάρων που υπερκφράζουν την τρανσγελίνη έλαβαν χώρα πειράματα διπλού ανοσοφθορισμού χρησιμοποιώντας δείκτες για κύτταρα του ανοσοποιητικού/ επιθηλιακά κύτταρα, για ενδοθηλιακά κύτταρα καθώς επίσης και για ενεργοποιημένους ινοβλάστες. Προκειμένου να βρεθεί ένας μηχανισμός μέσα από τον οποίο η τρανσγελίνη υπερεκφράζεται, προχωρήσαμε σε ανάλυση του υποκινητή της τρανσγελίνης. Τέλος, για να μελετήσουμε την πιθανότητα υπερέκφρασης της τρανσγελίνης σε νεφρικές ασθένειες, εξετάστηκε υλικό βιοψιών από αρκετούς ασθενείς (Ν=67) με ανοσοϊστοχημεία και διπλό ανοσοφθορισμό.Με Real-Time PCR, βρέθηκαν ότι δύο ισομορφές της τρανσγελίνης, αν και έχουν αυξημένη έκφραση στα ινωτικά ζώα, παρουσιάζουν ελαφρώς διαφορετικές κινητικές. Επιπλέον, παρατηρήθηκε ότι ήδη από τα πρώιμα στάδια της ίνωσης (2 μέρες), η τρανσγελίνη εντοπίζεται σε ένα μεγάλο ποσοστό στο διάμεσο χώρο και κυρίως σε κύτταρα που περιβάλλουν τα σπειράματα. Επιβεβαιώθηκε ότι τα κύτταρα που εκφράζουν την τρανσγελίνη είναι κύτταρα που μοιάζουν με ινοβλάστες, με βάση τον εκτεταμένο και σε υψηλό ποσοστό συνεντοπισμό της τρανσγελίνης με την αSMA, ενώ δεν είναι ενδοθηλιακά, επιθηλιακά και κύτταρα του ανοσοποιητικού συστήματος, βάση της έλλειψης συνεντοπισμού της τρανσγελίνης με το RECA-1, galectin 3 και το CD11b. Επιπλέον, δεν παρατηρήθηκε συνεντοπισμός της τρανσγελίνης με την S100A4/FSp1. Επιπρόσθετα, αυτοί οι περισπειραματικοί ινοβλάστες φαίνεται πως εκφράζουν την τρανσγελίνη κυτταροπλασματικά και πυρηνικά. Με την ανάλυση του υποκινητή της τρανσγελίνης, βρέθηκε ότι ανάμεσα σε αρκετούς μεταγραφικούς παράγοντες που έχουν ειδικά σημεία πρόσδεσης επάνω στον υποκινητή της τρανσγελίνης, μόνο o Kruppel-like factor 6 (KLF6) μεταγραφικός παράγοντας φαίνεται να συνεντοπίζεται με την τρανσγελίνη. Το εύρημα αυτό είναι σημαντικό για το μηχανισμό μέσα από το οποίο η τρανσγελίνη υπερεκφράζεται.Τέλος, τουλάχιστον σε 3 (από τις 9) κατηγορίες νεφρικών παθήσεων που μελετήθηκαν, την μεμβρανώδη νεφροπάθεια, την IgA νεφροπάθεια και την εστιακή τμηματική σπειραματοσκλήρυνση, η έκφραση της τρανσγελίνης εντοπίστηκε στα σπειράματα και σε κύτταρα γύρω από τα σπειράματα που μοιάζουν με ινοβλάστες, ενώ σε φυσιολογικό νεφρικό ιστό ανθρώπου, η τρανσγελίνη δεν εντοπίστηκε ποτέ σε αυτά τα σημεία.Συνοψίζοντας, τα δεδομένα μας για την τρανσγελίνη δείχνουν ότι η έκφραση της αυξάνεται σε ινωτικές νεφρικές ασθένειες και θα μπορούσε στο μέλλον να χρησιμοποιηθεί ως δείκτης για την νεφρική ίνωση.

scholarly journals SETDB1 promotes glioblastoma growth via CSF-1-dependent macrophage recruitment by activating the AKT/mTOR signaling pathway

2020 ◽  
Vol 39 (1) ◽  
Author(s):  
Shuai Han ◽  
Wei Zhen ◽  
Tongqi Guo ◽  
Jianjun Zou ◽  
Fuyong Li
Keyword(s):  
Cell Proliferation ◽  
Tumor Microenvironment ◽  
Real Time ◽  
Western Blotting ◽  
Real Time Pcr ◽  
Migration And Invasion ◽  
Common Disease ◽  
High Morbidity ◽  
Macrophage Recruitment ◽  
Syngeneic Mouse Model

Abstract Background Glioblastoma is a common disease of the central nervous system (CNS), with high morbidity and mortality. In the infiltrate in the tumor microenvironment, tumor-associated macrophages (TAMs) are abundant, which are important factors in glioblastoma progression. However, the exact details of TAMs in glioblastoma progression have yet to be determined. Methods The clinical relevance of SET domain bifurcated 1 (SETDB1) was analyzed by immunohistochemistry, real-time PCR and Western blotting of glioblastoma tissues. SETDB1-induced cell proliferation, migration and invasion were investigated by CCK-8 assay, colony formation assay, wound healing and Transwell assay. The relationship between SETDB1 and colony stimulating factor 1 (CSF-1), as well as TAMs recruitment was examined by Western blotting, real-time PCR and syngeneic mouse model. Results Our findings showed that SETDB1 upregulated in glioblastoma and relative to poor progression. Gain and loss of function approaches showed the SETDB1 overexpression promotes cell proliferation, migration and invasion in glioblastoma cells. However, knockdown SETDB1 exerted opposite effects in vitro. Moreover, SETDB1 promotes AKT/mTOR-dependent CSF-1 induction and secretion, which leads to macrophage recruitment in the tumor, resulted in tumor growth. Conclusion Our research clarified that SETDB1 regulates of tumor microenvironment and hence presents a potential therapeutic target for treating glioblastoma.

scholarly journals Expression and potential role of CXCL5 in the pathogenesis of intrauterine adhesions

2021 ◽  
Vol 49 (3) ◽  
pp. 030006052199771
Author(s):  
Zi-Ang Fang ◽  
Yu He ◽  
Chao Sun ◽  
Lei Zhan ◽  
Guiju Zhou ◽  
...  
Keyword(s):  
Real Time ◽  
Western Blotting ◽  
Rat Model ◽  
Real Time Pcr ◽  
Connective Tissues ◽  
Intrauterine Adhesions ◽  
Endometrial Cells ◽  
Protein Levels ◽  
Chemokine Ligand

Objective C-X-C motif chemokine ligand 5 (CXCL5), a member of the chemokine family, is associated with remodeling of connective tissues. However, its role in formation of intrauterine adhesions (IUA) remains unclear. We aimed to investigate the expression and mechanism underlying the role of CXCL5 in IUA. Methods Expression of CXCL5 in IUA was detected by immunohistochemistry in a rat model of IUA and by real-time PCR and western blotting in patients with IUA. The protein levels of matrix metalloproteinase 9 (MMP9) and transcription factor p65 in human endometrial cells were assessed by western blotting after CXCL5 overexpression. Results Protein expression of CXCL5 was significantly decreased in the endometria of IUA rats compared with that of control and sham-operated rats. Real-time PCR and western blotting in patients with IUA showed similar results to those from the rat model. After overexpression, CXCL5 significantly upregulated expression of MMP9 and slightly upregulated expression of p65 in human endometrial cells. Conclusions CXCL5 plays an important role in IUA formation after endometrial injury. We propose a molecular mechanism to explain formation of IUA, including downregulation of MMP9 by low CXCL5 expression. These findings provide valuable information for the prevention and targeted therapy of IUA.

scholarly journals Hypoxia and its possible relationship with endometrial receptivity in adenomyosis: a preliminary study

2021 ◽  
Vol 19 (1) ◽  
Author(s):  
Song Guo ◽  
Di Zhang ◽  
Xiaowei Lu ◽  
Qian Zhang ◽  
Ruihuan Gu ◽  
...  
Keyword(s):  
Real Time ◽  
Western Blotting ◽  
Real Time Pcr ◽  
Underlying Mechanism ◽  
Endometrial Receptivity ◽  
Control Group ◽  
Expression Levels ◽  
Preliminary Study

Abstract Background Adenomyosis (AM) is an important cause of female infertility. However, the underlying mechanism remains unclear. This report describes a preliminary study of hypoxia and its possible association with endometrial receptivity in AM. Methods The study was divided into in vitro and in vivo experiments. In vitro, expression levels of the endometrial receptivity markers HOXA10 and HOXA11 in the implantation period were examined using real-time PCR and western blotting. Endometrial expression of hypoxia-inducible factor (HIF)-1α, HIF-2α, and HIF-3α was determined using immunohistochemistry. In vivo, using an AM mouse model established by oral administration of tamoxifen, we inhibited expression of HIF-2α using an HIF-2α antagonist (PT2399; 30 mg/kg body weight, twice daily by oral gavage for 2 days) and then examined expression levels of Hoxa10 and Hoxa11 using real-time PCR and western blotting. Results Endometrial mRNA and protein expression levels of HOXA10 and HOXA11 were significantly lower in patients with AM than in control patients. Expression of HIF-2α was significantly higher in the AM group than in the control group, whereas that of HIF-1α and HIF-3α was equivalent in both groups. In vivo analysis showed that administration of the HIF-2α antagonist resulted in increased expression of Hoxa10 and Hoxa11 at both the mRNA and protein levels in AM model mice. Conclusions HIF-2α overexpression may be one reason for decreased endometrial receptivity in AM. The current findings provide insight into HIF-2α-mediated AM-related infertility and suggest that PT2399 has potential as a treatment for AM. Trial registration This trial was retrospectively registered.

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