Long-term follow-up of photodynamic therapy of cervical intraepithelial neoplasia grade 2 (CIN2).

BACKGROUND: To determine the long-term efficacy and safety of 5-aminolevulinic acid-based photodynamic therapy (ALA-PDT) for treating cervical intraepithelial neoplasia grade 2 (CIN2) as well as the suitability of ALA-PDT in treating of cervical lesions divided into cervical transformation zone type 3. METHODS: We included 81 patients diagnosed with CIN2 at the Department of Gynecology of the Affiliated Hospital of Qingdao University with data collected between January 2019 and January 2021 following ALA-PDT. Furthermore, we analyzed the superiority of ALA-PDT in fertility preservation among women of childbearing age based on follow-up data from 11 patients with fertility requirements. RESULTS: Our findings confirmed the long-term efficacy of ALA-PDT for CIN2 treatment, with an overall efficacy of 95.83 % (23/24) at follow-up of 25-36 months. Moreover, the cervical transformation zone type 3 improvement and human papillomavirus (HPV)-negative efficacy were 69.2 % (18/26) and 82.4 % (14/17), respectively. ALA-PDT is recommended for consenting patients with cervical transformation zone type 3. Additionally, women without primary infertility could experience natural pregnancy and full-term birth of more than one baby following ALA-PDT for CIN2 treatment, with a satisfaction rate of ≈100 %. CONCLUSIONS: ALA-PDT is recommendable for treating high-grade squamous intraepithelial lesions, especially in patients with fertility requirements.

RCAN family member 3 deficiency contributes to noncompaction of the ventricular myocardium.

Noncompaction of the ventricular myocardium (NVM), the third most diagnosed cardiomyopathy, is characterized by prominent trabeculae and intratrabecular recesses. However, the genetic etiology of 40-60% of NVM cases remains unknown. We identified two infants with NVM, in a nonconsanguineous family, with a typical clinical presentation of persistent bradycardia since the prenatal period. A homozygous missense variant (R223L) of RCAN family member 3 (RCAN3) was detected in both infants using whole-exome sequencing. In the zebrafish model, marked cardiac dysfunction was detected in rcan3 deficiency (MO-rcan3ATG-injected) and rcan-/- embryos. Developmental dysplasia of both endocardial and myocardial layers was also detected in rcan3-deficient embryos. RCAN3 R223L variant mRNAs did not rescue heart defects caused by rcan3 knockdown or knockout; however, hRCAN3 mRNA rescued these phenotypes. RNA-seq experiments showed that several genes involved in cardiomyopathies were significantly regulated through multiple signaling pathways in the rcan3-knockdown zebrafish model. In human cardiomyocytes, RCAN3 deficiency resulted in reduced proliferation and increased apoptosis, together with an abnormal mitochondrial ultrastructure. Thus, we suggest that RCAN3 is a susceptibility gene for cardiomyopathies, especially NVM and that the R223L mutation is a potential loss-of-function variant.

Loss of keratin 23 enhances growth inhibitory effect of melatonin in gastric cancer.

To investigate the effect of keratin 23 (KRT23) on the anticancer activity of melatonin (MLT) against gastric cancer (GC) cells, microarray analysis was applied to screen differentially expressed genes in AGS GC cells following MLT treatment. Western blotting was used to detect the expression of KRT23 in GC cells and normal gastric epithelial cell line GES­1. KRT23 knockout was achieved by CRISPR/Cas9. Assays of cell viability, colony formation, cell cycle, electric cell­substrate impedance sensing and western blotting were conducted to reveal the biological functions of KRT23­knockout cells without or with MLT treatment. Genes downregulated by MLT were enriched in purine metabolism, pyrimidine metabolism, genetic information processing and cell cycle pathway. Expression levels of KRT23 were downregulated by MLT treatment. Expression levels of KRT23 in AGS and SNU­216 GC cell lines were significantly higher compared with normal gastric epithelial cell line GES­1. KRT23 knockout led to reduced phosphorylation of ERK1/2 and p38, arrest of the cell cycle and inhibition of GC cell proliferation. Moreover, KRT23 knockout further enhanced the inhibitory activity of MLT on the tumor cell proliferation by inhibiting the phosphorylation of p38/ERK. KRT23 knockout contributes to the antitumor effects of MLT in GC via suppressing p38/ERK phosphorylation. In the future, KRT23 might be a potential prognostic biomarker and a novel molecular target for GC.

Novel bi-allelic variants of CHMP1A contribute to pontocerebellar hypoplasia type 8: additional clinical and genetic evidence.

Pontocerebellar hypoplasia type 8(PCH8) is a rare neurodegenerative disorder, reportedly caused by pathogenic variants of the CHMP1A in autosomal recessive inheritance, and CHMP1A variants have also been implicated in other diseases, and yet none of the prenatal fetal features were reported in PCH8. In this study, we investigated the phenotype and genotype in a human subject with global developmental delay, including clinical data from the prenatal stage through early childhood. Prenatally, the mother had polyhydramnios, and the bilateral ventricles of the fetus were slightly widened. Postnatally, the infant was observed to have severely delayed psychomotor development and was incapable of visual tracking before 2 years old and could not fix on small objects. The young child had hypotonia, increased knee tendon reflex, as well as skeletal malformations, and dental crowding; she also had severe and recurrent pulmonary infections. Magnetic resonance imaging of the brain revealed a severe reduction of the cerebellum (vermis and hemispheres) and a thin corpus callosum. Through whole exome sequencing and whole genomics sequencing, we identified two novel compound heterozygous variations in CHMP1A [c.53 T > C(p.Leu18Pro)(NM_002768.5) and exon 1 deletion region (NC_000016.10:g.89656392_89674382del)]. cDNA analysis showed that the exon1 deletion region led to the impaired expression, and functional verification with zebrafish embryos using base edition indicated variant c.53 T > C (p.Leu18Pro), causing dysplasia of the cerebellum and pons. These results provide further evidence that CHMP1A variants in a recessive inheritance pattern contribute to the clinical characteristics of PCH8 and further expand our knowledge of the phenotype and genotype spectrum of PCH8.

Embryonic organizer formation disorder leads to multiorgan dysplasia in Down syndrome.

Despite the high prevalence of Down syndrome (DS) and early identification of the cause (trisomy 21), its molecular pathogenesis has been poorly understood and specific treatments have consequently been practically unavailable. A number of medical conditions throughout the body associated with DS have prompted us to investigate its molecular etiology from the viewpoint of the embryonic organizer, which can steer the development of surrounding cells into specific organs and tissues. We established a DS zebrafish model by overexpressing the human DYRK1A gene, a highly haploinsufficient gene located at the "critical region" within 21q22. We found that both embryonic organizer and body axis were significantly impaired during early embryogenesis, producing abnormalities of the nervous, heart, visceral, and blood systems, similar to those observed with DS. Quantitative phosphoproteome analysis and related assays demonstrated that the DYRK1A-overexpressed zebrafish embryos had anomalous phosphorylation of ß-catenin and Hsp90ab1, resulting in Wnt signaling enhancement and TGF-ß inhibition. We found an uncovered ectopic molecular mechanism present in amniocytes from fetuses diagnosed with DS and isolated hematopoietic stem cells (HSCs) of DS patients. Importantly, the abnormal proliferation of DS HSCs could be recovered by switching the balance between Wnt and TGF-ß signaling in vitro. Our findings provide a novel molecular pathogenic mechanism in which ectopic Wnt and TGF-ß lead to DS physical dysplasia, suggesting potential targeted therapies for DS.

Sex Differences in Protein Expression and Their Perturbations in Amniotic Fluid Cells of Down Syndrome Fetuses.

Down syndrome (DS) is the most common chromosomal condition associated with intellectual disability and is characterized by a variety of additional clinical findings. The pathogenesis of DS and the differences between the sexes are not clear. In order to identify differentially expressed proteins that might be employed as potential biological markers and elucidate the difference in pathogenesis between different genders of T21 fetuses, providing clues for individualized detection and treatment is essential. Amniocyte samples of T21 males, T21 females, CN males, and CN females were collected by amniocentesis. The quantitative value of the peptide corresponding to each sample was determined through quantitative analysis by mass spectrometry. We identified many differentially expressed proteins between T21 fetuses and CN fetuses/T21 males and CN males/T21 females and CN females/and T21 males and T21 females. These differential proteins are associated with many important biological processes and affect the development of multiple systems, including the heart, hematopoietic, immune, reproductive, and nervous systems. Our results show sex-specific modulation of protein expression and biological processes and provide new insights into sex-specific differences in the pathogenesis of DS.

High cystic fibrosis transmembrane conductance regulator expression in childhood B-cell acute lymphoblastic leukemia acts as a potential therapeutic target.

Background: The role of cystic fibrosis transmembrane conductance regulator (CFTR) in hematopoiesis and adult leukemia has been demonstrated using a zebrafish model and leukemia cell lines in our previous works. Here, we continue to explore the association between CFTR and human childhood B-cell acute lymphoblastic leukemia (B-ALL). Methods: We continued to collect the peripheral blood and bone marrows of human childhood patients diagnosed with primary B-ALL as well as non-leukemia controls and isolated lymphocytes for analysis using western blotting and quantitative real-time polymerase chain reaction (qPCR) assay. Then, we used immunofluorescence, co-immunoprecipitation, western blotting, luciferase, 3-(4,5)-dimethylthiahiazo (-z-y1)-3,5-di-phenytetrazoliumromide (MTT) assays to identify the interaction of CFTR with Wnt signaling in B-ALL. Finally, we established B-ALL xenograft model in non-obese diabetic/severe combined immunodeficiency (NOD/SCID) mice using SUP-B15 cells, and examined whether the CFTR inhibitor CFTR-inh172 could active against SUP-B15-Dependent B-ALL in vivo. Results: Highly expressed CFTR protein and mRNA are associated with primary childhood B-ALL patients. Aberrantly upregulated CFTR and Wnt signaling, our previously reported CFTR-Dvl2-ß-catenin pathway, is found in human childhood B-ALL patients. Interference with CFTR in B-ALL cell lines induces the downregulation of DVL2/ß-catenin and Wnt downstream target accompanied by a reduction of cell proliferation. Furthermore, B-ALL cell lines SUP-B15 cell-transplanted NOD/SCID mice treated with CFTR inhibitor CFTRinh-172 had significantly longer survival and slower leukemia progression compared with mice treated with vehicle dimethyl sulfoxide (DMSO). Conclusions: These findings demonstrate that highly expressed CFTR is associated with human childhood B-ALL and the potential of CFTR inhibitor CFTR-inh172 for the treatment of human B-ALL.

Long non-coding RNA PTAR inhibits apoptosis but promotes proliferation, invasion and migration of cervical cancer cells by binding miR-101.

In this study, the expression of PTAR in cervical cancer tissues and cells was quantified by real-time PCR. Then, the roles of PTAR in HeLa cell proliferation and cell cycle were analyzed by a CCK-8 assay and flow cytometry, respectively.The effects of PTAR on cell migration and invasion were checked by Transwell and wound healing assays.The effect of PTAR on HeLa cell apoptosis was analyzed using annexin V/FITC staining. Finally, the interaction between PTAR and miR-101 in uterine cancer was verified through a dual-luciferase reporter assay and correlation analysis. The results showed that PTAR expression was aberrantly ascended in cervical cancer tissues and cell lines (Caski, SW756, SiHa, C33A and HeLa cells). Overexpressed PTAR could promote cell proliferation, migration and invasion in HeLa cells, which were suppressed by PTAR knockdown. Moreover, cell cycle progression stalled at the G1-G0 phase could be released with PTAR overexpression. The transfection of a PTAR vector inhibited apoptosis, while si-PTAR transfection increased apoptosis. Furthermore, PTAR could act as an endogenous sponge by directly binding to miR-101 and downregulating miR-101 expression. In conclusion, lncRNAPTAR plays a vital role and may be an effective target for the diagnosis and therapy of cervical cancer.

CFTR regulates embryonic T lymphopoiesis via Wnt signaling in zebrafish.

The number and function of T cells are abnormal as observed in cystic fibrosis (CF) patients and CF mouse models, and our previous work shows that the CFTR mutant leads to deficiency of primitive and definitive hematopoietic in zebrafish. However, the functions and underlying mechanisms of CFTR in T cell development during early embryogenesis have not been explored. Here, we report that the genetic ablation of CFTR in zebrafish resulted in abrogated embryonic T lymphopoiesis, which was ascribed to impaired thymic homing and expansion of hematopoietic stem cells (HSCs). Transcriptome analysis of isolated HSCs in zebrafish embryos at 48 hpf showed a significant alteration of key factors essential for T cell development and Wnt signaling, consistent with our previous work on CFTR regulating hematopoiesis. In brief, we uncovered the function of CFTR in embryonic T cell development and suggest that the immune deficiency of CF patients may originate from an early embryonic stage.

Integrative Pan-Cancer Analysis Reveals Decreased Melatonergic Gene Expression in Carcinogenesis and RORA as a Prognostic Marker for Hepatocellular Carcinoma.

BACKGROUND: Melatonin has been shown to play a protective role in the development and progression of cancer. However, the relationship between alterations in the melatonergic microenvironment and cancer development has remained unclear. METHODS: We performed a comprehensive investigation on 12 melatonergic genes and their relevance to cancer occurrence, progression and survival by integrating multi-omics data from microarray analysis and RNA sequencing across 11 cancer types. Specifically, the 12 melatonergic genes that we investigated, which reflect the melatonergic microenvironment, included three membrane receptor genes, three nuclear receptor genes, two intracellular receptor genes, one synthetic gene, and three metabolic genes. RESULTS: Widely coherent underexpression of nuclear receptor genes, intracellular receptor genes, and metabolic genes was observed in cancerous samples from multiple cancer types compared to that in normal samples. Furthermore, genomic and/or epigenetic alterations partially contributed to these abnormal expression patterns in cancerous samples. Moreover, the majority of melatonergic genes had significant prognostic effects in predicting overall survival. Nevertheless, few corresponding alterations in expression were observed during cancer progression, and alterations in expression patterns varied greatly across cancer types. However, the association of melatonergic genes with one specific cancer type, hepatocellular carcinoma, identified RORA as a tumor suppressor and a prognostic marker for patients with hepatocellular carcinoma. CONCLUSIONS: Overall, our study revealed decreased melatonergic gene expression in various cancers, which may help to better elucidate the relationship between melatonin and cancer development. Taken together, our findings highlight the potential prognostic significance of melatonergic genes in various cancers.

Cystic fibrosis transmembrane conductance regulator (CFTR) regulates embryonic organizer formation during zebrafish early embryogenesis.

Cystic fibrosis (CF) is associated with the manifestation of a number of medical conditions throughout the body. This prompted us to investigate the etiology of CF from the viewpoint of the embryonic organizer, which is responsible for steering the movement of surrounding cells into specific organs and tissues. In our previous work, we found that a cftr mutant had decreased nuclear ß-catenin levels in the early embryo at 5 hours post-fertilization (hpf), when the organizer forms. It is known that nuclear ß-catenin signaling is essential for the induction of the dorsal organizer. Therefore, we explored the role of cftr in the formation of the embryonic organizer in this work. Indeed, the expression of organizer and germ layer markers was significantly affected in cftr mutant embryos dependent on Wnt/ß-catenin signaling. Furthermore, quantitative proteome analysis revealed that the cftr mutant induced significant alteration in the expression of proteins related to many critical biological processes, cellular components, molecular functions, and signaling pathways, except for the Wnt/ß-catenin pathway. These findings demonstrate the function of cftr in embryonic organizer formation and provide an explanation for why many abnormalities occur in the bodies of CF patients.

Transcriptomic analysis on the effects of melatonin in gastrointestinal carcinomas.

BACKGROUND: Melatonin has been shown with anticancer property and therapeutic potential for tumors. However, there lacks a systematic study on the molecular pathways of melatonin and its antitumor effects in gastrointestinal carcinomas. METHODS: Using the gene expression profiles of four cancer cell lines from three types of gastrointestinal carcinomas before and after melatonin treatment, including gastric carcinoma (GC), colorectal carcinoma (CRC) and hepatocellular carcinoma (HCC), differentially expressed genes (DEGs) and biological pathways influenced by melatonin were identified. The qRT-PCR analyses were performed to validate the effects of melatonin on 5-FU resistance-related genes in CRC. RESULTS: There were 17 pathways commonly altered by melatonin in the three cancer types, including FoxO signaling pathways enriched by the upregulated DEGs and cell cycle signaling pathways enriched by the downregulated DEGs, confirmed the dual role of melatonin to tumor growth, pro-apoptosis and anti-proliferation. DEGs upregulated in the three types of cancer tissues but reversely downregulated by melatonin were commonly enriched in RNA transport, spliceosome and cell cycle signaling pathways, which indicate that melatonin might exert antitumor effects through these pathways. Our results further showed that melatonin can downregulate the expression levels of 5-FU resistance-related genes, such as thymidylate synthase in GC and ATR, CHEK1, BAX and MYC in CRC. The qRT-PCR results demonstrated that melatonin enhanced the sensitivity of CRC 5-FU resistant cells by decreasing the expression of ATR. CONCLUSIONS: Melatonin exerts the effects of pro-apoptosis and anti-proliferation on gastrointestinal carcinomas, and might increase the sensitivity of 5-FU in GC and CRC patients.

CFTR deficiency causes cardiac dysplasia during zebrafish embryogenesis and is associated with dilated cardiomyopathy.

Mutations in the CFTR gene cause cystic fibrosis (CF) with myocardial dysfunction. However, it remains unknown whether CF-related heart disease is a secondary effect of pulmonary disease, or an intrinsic primary defect in the heart. Here, we used zebrafish, which lack lung tissue, to investigate the role of CFTR in cardiogenesis. Our findings demonstrated that the loss of CFTR impairs cardiac development from the cardiac progenitor stage, resulting in cardiac looping defects, a dilated atrium, pericardial edema, and a decrease in heart rate. Furthermore, we found that cardiac development was perturbed in wild-type embryos treated with a gating-specific CFTR channel inhibitor, CFTRinh-172, at the blastula stage of development, but not at later stages. Gene expression analysis of blastulas indicated that transcript levels, including mRNAs associated with cardiovascular diseases, were significantly altered in embryos derived from cftr mutants relative to controls. To evaluate the role of CFTR in human heart failure, we performed a genetic association study on individuals with dilated cardiomyopathy and found that the I556V mutation in CFTR, which causes a channel defect, was associated with the disease. Similar to other well-studied channel-defective CFTR mutants, CFTR I556V mRNA failed to restore cardiac dysplasia in mutant embryos. The present study revealed an important role for the CFTR ion channel in regulating cardiac development during early embryogenesis, supporting the hypothesis that CF-related heart disease results from an intrinsic primary defect in the heart.

Application of 5-aminolevulinic acid photodynamic therapy for vaginal intraepithelial neoplasia, a report of six cases.

BACKGROUND: Vaginal intraepithelial neoplasia (VAIN) has a high-risk for recurrence and may precede genital cancers, such as vaginal cancer and/or other invasive diseases. Human papillomavirus (HPV)-induced VAIN may occur after loop electrosurgical excision procedures (LEEPs) or panhysterectomy. 5-aminolevulinic acid (ALA) photodynamic therapy (ALA-PDT) has demonstrated utility in preventing the recurrence of cervical intraepithelial neoplasia (CIN); however, evaluation of its effect on VAIN has not been performed. METHODS: The effectiveness of ALA-PDT was evaluated in 6 women diagnosed with HPV-induced VAIN. Lesion treatment was performed 3 h after ALA using light at a wavelength of 635 nm and light density of 80 mw/cm2. Therapeutic effect was assessed using HPV-DNA tests combined with liquid-based cervical cytology (LCT). RESULTS: Six women, aged 49-54 years, who were diagnosed VAIN grade 1/2 or 2 after LEEP or panhysterectomy or no surgery underwent ALA-PDT (range, 4-8 treatments). Four of the 6 women were HPV negative on retesting 3-4 months after ALA-PDT. Most patients exhibited no signs of recurrence during the follow-up period. CONCLUSIONS: Direct use of ALA-PDT or after panhysterectomy did not necessarily lead to a negative result; however, ALA-PDT after LEEP or panhysterectomy combined with LEEP yielded a satisfactory curative effect on VAIN. Although recurrence rates need to be monitored in longer-term studies, the absence of post-treatment complications in this study supports the potential utility of the technique.

A qualitative transcriptional prognostic signature for patients with stage I-II pancreatic ductal adenocarcinoma.

Accurately prognostic evaluation of patients with stage I-II pancreatic ductal adenocarcinoma (PDAC) is of importance to treatment decision and patient management. Most previously reported prognostic signatures were based on risk scores summarized from quantitative expression measurements of signature genes, which are susceptible to experimental batch effects and impractical for clinical applications. Based on the within-sample relative expression orderings of genes, we developed a robust qualitative transcriptional prognostic signature, consisting of 64 gene pairs (64-GPS), to predict the overall survival (OS) of 161 stage I-II PDAC patients in the training dataset who were treated with surgery only. Samples were classified into the high-risk group when at least 25 of 64 gene pairs suggested it was at high risk. The signature was successfully validated in 324 samples from 6 independent datasets produced by different laboratories. All samples in the low-risk group had significantly better OS than samples in the high-risk group. Multivariate Cox regression analyses showed that the 64-GPS remained significantly associated with the OS of patients after adjusting available clinical factors. Transcriptomic analysis of the 2 prognostic subgroups showed that the differential expression signals were highly reproducible in all datasets, whereas the differences between samples grouped by the TNM staging system were weak and irreproducible. The epigenomic analysis showed that the epigenetic alternations may cause consistently transcriptional changes between the 2 different prognostic groups. The genomic analysis revealed that mutation­induced disturbances in several key genes, such as LRMDA, MAPK10, and CREBBP, might lead to poor prognosis for PDAC patients. Conclusively, the 64-GPS can robustly predict the prognosis of patients with stage I-II PDAC, which provides theoretical basis for clinical individualized treatment.

Effect of melatonin on T/B cell activation and immune regulation in pinealectomy mice.

Melatonin is an indole neuroendocrine hormone that is mainly secreted by the pineal gland to regulate circadian rhythm, antioxidation, and immune regulation. Melatonin plays an important role in T cell-mediated immune responses against cancer, infections, and the development of many autoimmune diseases. The aim of this study was to investigate the immunomodulatory effects of melatonin on T/B cell activation in pinealectomy mice. The improved pinealectomy procedure for mice presented in this study is a good animal model to be used in follow-up studies on melatonin. After pinealectomy, the tissue removed was identified as the pineal body using HE staining. The effects of melatonin supplementation on T cell activation and activation-related changes to the MAPK/NF-κ B pathways were analyzed by flow cytometry and real-time PCR. We found that expression levels of Th1, Th2 and Th17-related cytokines in peripheral blood were lower in mice that had undergone pinealectomy, compared with normal mice. After melatonin supplementation, cytokine levels rapidly increased within a short period of time, which resulted in the gradual recovery of cytokine expression levels. Moreover, activation of T/B cells in mice was weakened and decreased after pineal gland removal. Melatonin was found to inhibit the expression of TLR3, p38, JNK, and MAPK/NF-κ B within a short period (2 weeks) of melatonin replenishment. This inhibition gradually weakened with time, since the degree of inhibition is negatively related with the dosage of melatonin. In conclusion, melatonin may regulate the activation of T/B cells, playing a critical role in the regulation of immune balance.

Treatment of human T-cell acute lymphoblastic leukemia cells with CFTR inhibitor CFTRinh-172.

Our previous studies have demonstrated that a previously unrecognized role of CFTR in hematopoiesis and acute leukemia. Here, we show that CFTR inhibitor CFTR-inh172 possesses ability to inhibit human T-cell acute lymphoblastic leukemia cells. In detail, CFTR-inh172 inhibited cell proliferation, promoted apoptosis and arrested the cell cycle in human T-cell acute lymphoblastic leukemia cell CCRF-CEM, JURKAT and MOLT-4. Furthermore, transcriptome analysis reveals that CFTR-inh172 induces significant alteration of gene expression related to apoptosis and proliferation. These findings demonstrate the potential of CFTR inhibitor CFTR-inh172 in human T-cell acute lymphoblastic leukemia treatment.

Effect of emodin on T cell subsets in NOD mice with NaI­induced experimental autoimmune thyroiditis.

Chronic lymphocytic thyroiditis (CLT), also known as Hashimoto's thyroiditis, is an autoimmune disease in which the thyroid gland is gradually destroyed. To date, only a limited number of agents can effectively suppress thyroiditis development in CLT patients. The aim of the current study was to investigate the protective effect of emodin on experimental autoimmune thyroiditis (EAT) in mice, which is considered an excellent model for CLT. NaI was used to induce the EAT model in non­obese diabetic (NOD) mice. An ELISA method was employed to detect the TgAb level (thyroid inflammation) in the serum of the EAT mice. The T cell subsets in peripheral blood and spleen were detected by flow cytometry. The histopathological study revealed that the thyroid inflammatory cell infiltration was significantly reduced by emodin compared with the model group. In addition, ELISA assays indicated that the NaI­induced serum TgAb upregulation was dramatically revered by emodin. Moreover, the level of serum IFN­Î³ and the cell populations of CD3+CD4+IL­4+, CD3+CD4+ IFN­Î³+, CD3+CD8+IL­4+, CD3+CD8+ IFN­Î³+ T cells in peripheral blood monocytes and splenic lymphocytes were significantly increased by NaI in the model group compared with in the normal group. Nevertheless, this type of increase was markedly attenuated by emodin. To conclude, the EAT model was successfully established by treating NOD mice with NaI. Emodin indicated an inhibitory effect on the autoimmune response that was significantly different in EAT compared with control mice. Furthermore, the anti­inflammatory action of emodin on EAT mice may be mediated via the inhibition of the secretion of IFN­Î³ and the cell numbers of CD3+CD4+IL­4+, CD3+CD4+ IFN­Î³+, CD3+CD8+IL­4+ and CD3+CD8+ IFN­Î³+ T cells in the peripheral blood monocytes and splenic lymphocytes. Therefore, the data may offer valuable insight on the efficacy of treatment of CLT with emodin.

CFTR is required for the migration of primordial germ cells during zebrafish early embryogenesis.

Mutations in the cystic fibrosis transmembrane conductance regulator (CFTR) gene affect fertility in both sexes. However, the involvement of CFTR in regulating germ cell development remains largely unknown. Here, we used zebrafish model to investigate the role of CFTR in primordial germ cells (PGCs) development. We generated a cftr frameshift mutant zebrafish line using CRISPR/Cas9 technique and investigated the migration of PGCs during early embryo development. Our results showed that loss of Cftr impairs the migration of PGCs from dome stages onward. The migration of PGCs was also perturbed by treatment of CFTRinh-172, a gating-specific CFTR channel inhibitor. Moreover, defected PGCs migration in cftr mutant embryos can be partially rescued by injection of WT but not other channel-defective mutant cftr mRNAs. Finally, we observed the elevation of cxcr4b, cxcl12a, rgs14a and ca15b, key factors involved in zebrafish PGCs migration, in cftr-mutant zebrafish embryos. Taken together, the present study revealed an important role of CFTR acting as an ion channel in regulating PGCs migration during early embryogenesis. Defect of which may impair germ cell development through elevation of key factors involved in cell motility and response to chemotactic gradient in PGCs.