T-box Transcription Factor Research Articles

Single-cell morphometrics reveals T-box gene-dependent patterns of epithelial tension in the Second Heart field

The vertebrate heart tube extends by progressive addition of epithelial second heart field (SHF) progenitor cells from the dorsal pericardial wall. The interplay between epithelial mechanics and genetic mechanisms during SHF deployment is unknown. Here, we present a quantitative single-cell morphometric analysis of SHF cells during heart tube extension, including force inference analysis of epithelial stress. Joint spatial Principal Component Analysis reveals that cell orientation and stress direction are the main parameters defining apical cell morphology and distinguishes cells adjacent to the arterial and venous poles. Cell shape and mechanical forces display a dynamic relationship during heart tube formation. Moreover, while the T-box transcription factor Tbx1 is necessary for cell orientation towards the arterial pole, activation of Tbx5 in the posterior SHF correlates with the establishment of epithelial stress and SHF deletion of Tbx5 relaxes the progenitor epithelium. Integrating findings from cell-scale feature patterning and mechanical stress provides new insights into cardiac morphogenesis.

Transcriptional regulation of olfactory receptor OR51B5 by the TBX6.

Olfactory receptors (ORs) are G protein-coupled receptors primarily expressed in olfactory tissue, facilitating the perception of odors. Interestingly, they have also been detected in non-olfactory tissues such as the skin, where they regulate processes like collagen synthesis. This study aimed to analyze the promoter of the OR family 51 subfamily B member 5 (OR51B5) and identify the transcription factors that bind to it to understand the potential regulatory mechanisms for OR51B5 expression. We examined the promoter region spanning 2,000 base pairs upstream of the transcription start site and conducted a deletion analysis, revealing that the core promoter encompasses the region from -153 to -111 base pairs. A luciferase assay using various candidate transcription factors showed that the overexpression or knockdown of T-Box Transcription Factor 6 (TBX6) significantly regulated OR51B5 promoter activity, while other candidate transcription factors had no significant effect. Additionally, we validated TBX6 binding to the OR51B5 promoter using site-directed mutation and electrophoretic mobility shift assays. This study is the first to uncover the role of TBX transcription factors in regulating OR gene expression in mammals, which may have implications for treating related disorders.

Shifts in embryonic oxygen levels cue heterochrony in limb initiation.

Heterochrony, or the alteration of developmental timing, is an important mechanism of evolutionary change. Avian species display synchronized growth of the forelimbs and hindlimbs, while mammalian species show delayed hindlimb development. We find that mammalian limb heterochrony is evident from the start of limb bud formation, and is associated with heterochronic expression of T-box transcription factors. This heterochronic shift is not due to changes in cis-regulatory sequences controlling T-box gene expression, but unexpectedly, is dependent upon differential oxygen levels to which avian and mammalian embryos are exposed prior to limb initiation, mediated, at least partially, by an NFKB transcription factor, cRel. Together, these results provide mechanistic understanding of an important example of developmental heterochrony and exemplify how the maternal environment regulates timing during embryonic development.

6611 ACTH deficiency due to TBX19 mutation mimics complete adrenal insufficiency in a newborn

Abstract Disclosure: T. Reinehr: None. P.M. Holterhus: None. S.A. Wudy: None. A. Richter-Unruh: None. C. Spratte: None. C. Roll: None. Isolated ACTH deficiency (IAD) is a rare cause of adrenocortical insufficiency. T-box transcription factor 19 (TBX19) mutations are responsible for more than 60% of neonatal cases of IAD. To the best of our knowledge, we present for the first time a newborn with ACTH deficiency due to a TBX19 mutation with almost steroid-free plasma and urine: At day 2 of life, the female newborn (birth weight 3190g, length 52 cm, 40+1 gestational age) was transferred to our neonatal intensive care unit for severe hypoglycemia (12 mg/dl), which responded to intravenous glucose infusion. Surprisingly, the sodium serum concentrations decreased from 140 mmol/l at day 2 of life to 126 mmol/l at day 7 of life. There were no clinical signs of adrenal insufficiency such as vomiting or hypotonia. Treatment with oral sodium-chloride 5.85% was started. Endocrine diagnostic work-up performed at day 8 of life demonstrated normal values of fT4, HGH and IGFBP-3 ruling out pituitary involvement of these axes, while ACTH serum concentrations were markedly decreased (1.5 pg/ml). Liquid- chromatography-mass spectrometry (LC-MS/MS) revealed nearly absent glucocorticoids (17-hydroxyprogesterone <0.12 nmol/l, 11-deoxycortisol <0.08 nmol/l, cortisol <4.2 nmol/l, and cortisone 1.3 nmol/l (norm value [nv]: 17-138 nmol/l)). Androgens were undetectable (testosterone <0.02 nmol/l, androstenedione <0.08 nmol/l). Mineralocorticoids were severely reduced (11-desoxycorticosterone 0.1 nmol/l (nv: 0.2-1.0 nmol/l), corticosterone 0.6 nmol/l (nv: 1-24 nmol/l), aldosterone 0.1 nmol/l (nv: 0.8-2.3 nmol/l)). In urine spot, gas chromatography-mass spectrometry (GC-MS) demonstrated subnormal excretion of C19 and C21 steroids including fetal zone steroids. Serum and urine measurements were interpreted by the labs as a typical finding similar to StAR defect. The genetic analyses revealed a normal female karyotype (46,XX) and ruled out CYP11A1 and StAR, mutations but demonstrated a homozygous missense mutation in TBX19 (c.172A>G, p.(Thr58Ala)). After 3 months of life under treatment with hydrocortisone (10mg/m² divided in 3 doses) and fludrocortisone (0.05 mg divided in 2 doses), cortisol was normal (320 nmol/l) and DHEAS was undetectable (<3 nmol/l). While androgens were in normal range (testosterone 2.6 nmol/l, androstenedione 0.28 nmol/l), 11-deoxycorticosterone was normal (0.74 nmol/l), corticosterone was undetectable (<0.5 nmol/l) and aldosterone was decreased (0.12 nmol/l nv: 0.8-1.2 nmol/l). At this time point, ACTH was <4 pg/ml, renin 1.6 mIU/l, LH 0.7 mIU/l and FSH 24.5 mIU/ml. This case demonstrated that isolated ACTH deficiency should be considered as a potential differential diagnosis in complete adrenal steroid insufficiency in newborns. Our data suggest that ACTH is not only necessary for cortisol biosynthesis but also for sufficient mineralocorticoid and androgen production by the adrenals in neonates. Presentation: 6/3/2024

Circ_0060927 promotes colorectal cancer development by sponging miR-331-3p and upregulating TBX2

BackgroundThe dysregulation of circular RNAs (circRNAs) is closely associated with the pathogenesis of colorectal cancer (CRC). The present study aimed to elucidate the biological function and mechanism of circ_0060927 in CRC. Methods5-ethynyl-2’-deoxyuridine, Cell Counting Kit-8 (CCK-8), flow cytometry and transwell assays, as well as Xenograft tumor models were adopted for in vitro and in vivo analyses. The interaction between microRNA-331-3p (miR-331-3p) and circ_0060927 or T-box transcription factor 2 (TBX2) was verified by the dual-luciferase reporter and RNA pull-down assays. ResultsCirc_0060927 deficiency inhibited cell proliferation, autophagy, migration, and invasion and increased cell apoptosis and necrosis in CRC cells, as well as inhibited tumor growth in vivo. Circ_0060927 could bind to miR-331-3p, and circ_0060927 regulated CRC cell behaviors via sponging miR-331-3p. TBX2 was targeted by miR-331-3p, and miR-331-3p targeted TBX2 to exert the anti-cancer role in CRC cells. Mechanically, circ_0060927 regulated TBX2 expression by sequestering miR-331-3p in CRC cells. ConclusionCirc_0060927 downregulation inhibited CRC progression by regulating the miR-331-3p/TBX2 axis, which might offer a potential treatment target for CRC.

PTPN2 dephosphorylates STAT3 to ameliorate anesthesia-induced cognitive decline in aged rats by altering the microglial phenotype and inhibiting inflammation

Perioperative neurocognitive disorders (PNDs) are common neurological complications after anesthesia in the elderly. Protein tyrosine phosphatase non-receptor type 2 (PTPN2) regulates signal transducer and activator of transcription protein 3 (STAT3) signaling to control inflammation in certain organs, but its role in PNDs remains unknown. Herein, we constructed a PND model in 18-month-old rats by treating them with sevoflurane. PND rats developed neuroinflammation, along with a significant decrease in PTPN2 expression and a rise in STAT3 phosphorylation in the hippocampus. Ptpn2 overexpression alleviated the behavioral disorders of PND rats, ameliorated neuronal injury, inhibited neuroinflammation, inflammasome activation, microglial activation, and microglial phenotype switching. Similar results were observed in sevoflurane-treated HMC3 microglia with PTPN2 overexpression, while PTPN2 silencing showed the opposite results. Additionally, PTPN2 seems to be a target of T-box transcription factor 2 (TBX2). These results contribute to the evidence supporting the idea that PTPN2 is a regulatory factor in PND progression.

A High-Throughput Drug Repurposing Strategy to Treat TBX2 and/or TBX3 Dependent Cancers.

The highly homologous T-box transcription factors TBX2 and TBX3 are critical for embryonic development, and their overexpression in postnatal tissues contributes to a wide range of malignancies, including melanoma and rhabdomyosarcoma. Importantly, when TBX2 and TBX3 are depleted in cancers where they are overexpressed, the malignant phenotype is inhibited, and they have therefore been regarded as druggable targets. However, the time and costs associated with de novo drug development are challenging and result in drugs that are costly, especially for patients in low- and middle-income countries. In the current study, we therefore combined a targeted and drug repurposing approach to identify drugs that are expected to be more efficacious and cost-effective with significantly reduced side effects. A high-throughput cell-based immunofluorescence screen was performed to identify drugs in the Pharmakon 1600 drug library that can negatively regulate TBX2 and/or TBX3 levels. "Hit" drugs were validated for their effect on TBX2/TBX3 levels and cytotoxicity in TBX2/TBX3-dependent melanoma and rhabdomyosarcoma cells. To this end, immunofluorescence, western blotting, quantitative real-time PCR, and MTT cell viability assays were performed. Niclosamide, piroctone olamine, and pyrvinium pamoate, were identified as TBX2 and/or TBX3-targeting drugs, and they exhibited cytotoxicity in a TBX2/TBX3-dependent manner. Furthermore, these "Hit" drugs were shown to induce senescence and/or apoptosis. Niclosamide, piroctone olamine, and pyrvinium pamoate are promising, cost-effective therapeutic agents for the treatment of TBX2/TBX3-dependent cancers.

Identification of a T-box transcription factor TBX2 and its negative regulatory role in melanin production in the Pacific oyster Crassostrea gigas

Oyster shells exhibit varying color patterns—black, white, or black and white striations—attributable to differences in melanin content and distribution. In this study, we identified a new homolog of TBX2, a member of the T-box transcription factor family, in the Pacific oyster (Crassostrea gigas) named CgTBX2. The mRNA expression of CgTBX2 was higher in tissues from white-shelled oysters than in those from black-shelled oysters. Knockdown of CgTBX2 in C. gigas significantly upregulated the expression of two key genes involved in melanin synthesis, CgTYR and CgTYR2. The tissue expression profile revealed that CgTBX2 is highly expressed in gill and mantle. CgTBX2 protein is mainly localized in the nucleus and can combine with TBX-binding elements (TBEs) present in the CgOCA2 promoter. CgTBX2 repressed the activity of the CgOCA2 promoter, with a strong transcriptional inhibition effect on the −2000 to −878 and −211 to −1 regions. Furthermore, the combination of CgTBX2 with GTTGACACCTT sequence is responsible for the transcriptional inhibition on the −211 to −1 region of the CgOCA2 promoter. Our findings reveal that CgTBX2, a novel regulator of melanin pigmentation in C. gigas, negatively regulates melanin deposition by inhibiting tyrosinase genes expression and repressing the transcriptional activity of the CgOCA2 promoter.

T-Box Transcription Factor 2 Mediates Chemoresistance of Endometrial Cancer via Regulating FSP1-involved Ferroptosis.

Chemotherapy is increasingly being used in the first-line treatment of endometrial cancer (EC) patients. However, chemoresistance seriously affects its efficacy. Understanding the underlying molecular mechanisms is critical for EC treatment. We explored the regulatory role of T-Box transcription factor 2 (TBX2)-ferroptosis suppressor protein 1 (FSP1) axis in ferroptosis and chemoresistance of EC. Cisplatin-resistant cell line Ishikawa/DDP cells were utilized to generate TBX2 and FSP1 overexpression and knockdown stable cell lines by using lentivirus infection and puromycin selection. Cell viability and ferroptosis status were evaluated in EC cells with or without Cisplatin and/or FSP1 inhibitor (iFSP1) using CKK-8, lipid peroxidation, malondialdehyde, and lactate dehydrogenase release assays. Endometrial carcinoma xenograft mouse model was established to further explore the function of TBX2-FSP1 axis on ferroptosis and tumor progression in EC. TBX2 suppressed Cisplatin-induced ferroptosis through up-regulating FSP1 expression level in EC cells. On the contrary, knockdown of TBX2 reduced FSP1 expression and significantly promoted Cisplatin-induced ferroptosis. TBX2 or FSP1 overexpression and knockdown promote and inhibit EC tumor growth under Cisplatin treatment, respectively. Interestingly, silence FSP1 could reverse TBX2-mediated ferroptosis inhibition and tumor-promoting effect. TBX2-FSP1 axis inhibits ferroptosis and enhances the Cisplatin resistance, which will provide an important theoretical basis and potential solution for the clinical treatment of EC.

PCSK1N as a tumor size marker and an ER stress response protein in corticotroph pituitary adenomas.

Silent corticotroph adenoma (SCA) exhibits more tumor aggressiveness features than functioning adenomas (FCA). We aimed to investigate PCSK1N expression in CA and examine if ER stress-induced responses affect cell survival in a corticotroph tumor cell model. Clinical and imaging characteristics were recorded in 33 patients with FCA (20 women, 11 macroadenomas) and 18 SCA (8 women, all macroadenomas). Gene expression of proopiomelanocortin (POMC), T-box transcription factor 19(TBX19)/TPIT, proprotein convertase subtilisin/kexin type 1(PCSK1)/PC1/3, and its inhibitor PCSK1N, was measured by RT-qPCR in adenoma tissue.Mouse pituitary corticotroph tumor (AtT-20) cells were treated with tanespimycin (17-AAG), a HSP90 chaperone inhibitor, to induce ER stress, followed by gene and protein analyses. POMC, TPIT, and PCSK1 expression were higher, whereas PCSK1N was lower in FCA compared to SCA. PCSK1N correlated with POMC (rs= -0.514, p <0.001), TPIT (rs= -0.386, p = 0.005), PCSK1 (rs= -0.3691, p = 0.008), and tumor largest diameter (rs= 0.645, p <0.001), in all CA. Induction of ER stress by 17-AAG in AtT-20 cells led to a decrease of POMC and an increase of PCSK1N gene expression at 24h. Moreover, a downregulation of cell cycle, apoptosis, and senescence pathways, and alterations in cell adhesion and cytoskeleton were observed at the protein level. PCSK1N is higher in SCA compared with FCA, and associated with corticotroph cell markers and tumor size. PCSK1N is likely to be part of the adaptive response to ER stress, potentially conferring a survival advantage to the corticotroph tumor cell in conjunction with other proteins.

C-Myc, AKT, Hsc70, and the T-Box Transcription Factor TBX3 Form an Important Oncogenic Signaling Axis in Breast Cancer.

Breast cancer is the second leading cause of death in women globally, and it remains a health burden due to poor therapy response, cancer cell drug resistance, and the debilitating side effects associated with most therapies. One approach to addressing the need to improve breast cancer therapies has been to elucidate the mechanism(s) underpinning this disease to identify key drivers that can be targeted in molecular therapies. The T-box transcription factor, TBX3, is upregulated in breast cancer, in which it contributes to important oncogenic processes, and it has been validated as a potential therapeutic target. Here, we investigated the molecular mechanisms that upregulate TBX3 in breast cancer, and we show that it involves transcriptional activation by c-Myc, post-translational modification by AKT1 and AKT3, and interaction with the molecular chaperone Hsc70. Together, the results from this study provide evidence that c-Myc, AKT, Hsc70, and TBX3 form part of an important oncogenic pathway in breast cancer and thus reveal versatile ways of interfering with the oncogenic activity of TBX3 for the treatment of this neoplasm. Implications: Targeting the c-Myc/AKT/TBX3/Hsc70 signaling axis may be an effective treatment strategy for TBX3-driven breast cancer.

450. TISSUE RESIDENT MEMORY CELLS: CORRELATING THE PHENOTYPIC AND FUNCTIONAL CAPABILITIES OF A POTENTIAL TUMOUR REACTIVE CELL POPULATION IN OESOPHAGEAL ADENOCARCINOMA

Abstract Background Oesophageal adenocarcinoma (OAC) is a disease with a high mortality and morbidity rate. Immunotherapy has recently demonstrated progress in the treatment of OAC but its impact on long term survival is currently uncertain. Tissue resident memory (TRM) T cells are a subset of lymphocytes within the tumour/tissue infiltrating lymphocyte (TIL) milieu that has attracted considerable interest within cancer immunology. Although TRM cells have been associated with better outcomes in multiple cancer types there is a lack of knowledge of their role in OAC. This study aims to critically correlate the phenotype of TRM with their functional capacity in this disease. Methods 33 patients undergoing surgical resection for OAC were recruited and consented for collection of fresh tumour (T), adjacent normal tissue (AN), and peripheral blood. Peripheral blood mononuclear cell (PBMC) from blood and infiltrating lymphocytes from T and AN were isolated using standard density gradient and enzymatic digestion protocols. Multiparametric flow cytometry was performed on 12 patients using matched PBMC, T and AN infiltrating lymphocytes for phenotypic analysis and 21 patients for functional analysis following cell stimulation assay. BD FACSymphony was used for data acquisition and data analysed on FlowJo version 10.10. Statistical analysis was carried out using GraphPad Prism. Results TRM (CD103+CD69+) dominated the CD8+ T cell population of T and AN TIL. Expression of checkpoint proteins PD1+ and CD39+, a marker of tumour reactivity, was identified on CD8+ TRM and CD8+ non TRM in TIL. The differentiation status of TRM within memory lineages demonstrated that TRM express a higher proportion of effector memory cells (CCR7-CD45RA) compared to those in blood. TRM cells also showed lower expression of the transcription factors EOMES, TBET and TCF compared to CD8+ cells within PBMC. Moreover, tumour CD8+ TRM populations had reduced ability to produce IL-2 following stimulation compared to adjacent normal TRM populations. Conclusion TRM make up a large proportion of CD8+ T cell within the tumour microenvironment (TME) of OAC. TRM generation is associated with the downregulation of T-Box transcription factors EOMES and TBET and expression of an effector memory phenotype with reduction of the stem-like transcription factor, TCF. High levels of CD39+ indicate a strong level of activation but the increased PD1 expression and reduced capability to produce IL2 indicate functional impairment suppressed within the TME. An opportunity may exist to reverse this with novel immunotherapeutic agents.

Coordinated Tbx3 / Tbx5 transcriptional control of the adult ventricular conduction system.

The cardiac conduction system (CCS) orchestrates the electrical impulses that enable coordinated contraction of the cardiac chambers. The T-box transcription factors TBX3 and TBX5 are required for cardiac conduction system development and associated with overlapping and distinct human cardiac conduction system diseases. We evaluated the coordinated role of Tbx3 and Tbx5 in the murine ventricular conduction system (VCS). We engineered a compound Tbx3:Tbx5 conditional knockout allele for both genes located in cis on mouse chromosome 5. Conditional deletion of both T-box transcriptional factors in the ventricular conduction system, using the VCS-specific Mink:Cre, caused loss of VCS function and molecular identity. Combined Tbx3 and Tbx5 deficiency in the adult VCS led to conduction defects, including prolonged PR and QRS intervals and elevated susceptibility to ventricular tachycardia. These electrophysiologic defects occurred prior to detectable alterations in cardiac contractility or histologic morphology, indicative of a primary conduction system defect. Tbx3:Tbx5 double knockout VCS cardiomyocytes revealed a transcriptional shift towards non-CCS-specialized working myocardium, suggesting reprogramming of their cellular identity. Furthermore, optical mapping revealed a loss of VCS-specific conduction system propagation. Collectively, these findings indicate that Tbx3 and Tbx5 coordinate to control VCS molecular fate and function, with implications for understanding cardiac conduction disorders in humans.

Transcription factor 4 is a key mediator of oncogenesis in neuroblastoma by promoting MYC activity.

Super-enhancer-associated transcription factor networks define cell identity in neuroblastoma (NB). Dysregulation of these transcription factors contributes to the initiation and maintenance of NB by enforcing early developmental identity states. We report that the class I basic helix-loop-helix (bHLH) transcription factor 4 (TCF4; also known as E2-2) is a critical NB dependency gene that significantly contributes to these identity states through heterodimerization with cell-identity-specific bHLH transcription factors. Knockdown of TCF4 significantly induces apoptosis in vitro and inhibits tumorigenicity in vivo. We used genome-wide expression profiling, TCF4 chromatin immunoprecipitation sequencing (ChIP-seq) and TCF4 immunoprecipitation-mass spectrometry to determine the role of TCF4 in NB cells. Our results, along with recent findings in NB for the transcription factors T-box transcription factor TBX2, heart- and neural crest derivatives-expressed protein 2 (HAND2) and twist-related protein 1 (TWIST1), propose a role for TCF4 in regulating forkhead box protein M1 (FOXM1)/transcription factor E2F-driven gene regulatory networks that control cell cycle progression in cooperation with N-myc proto-oncogene protein (MYCN), TBX2, and the TCF4 dimerization partners HAND2 and TWIST1. Collectively, we showed that TCF4 promotes cell proliferation through direct transcriptional regulation of the c-MYC/MYCN oncogenic program that drives high-risk NB. Mechanistically, our data suggest the novel finding that TCF4 acts to support MYC activity by recruiting multiple factors known to regulate MYC function to sites of colocalization between critical NB transcription factors, TCF4 and MYC oncoproteins. Many of the TCF4-recruited factors are druggable, giving insight into potential therapies for high-risk NB. This study identifies a new function for class I bHLH transcription factors (e.g., TCF3, TCF4, and TCF12) that are important in cancer and development.

Tbxt alleviates senescence and apoptosis of nucleus pulposus cells through Atg7-mediated autophagy activation during intervertebral disk degeneration.

Intervertebral disk degeneration (IDD) is a significant cause of low back pain, characterized by excessive senescence and apoptosis of nucleus pulposus cells (NPCs). However, the precise mechanisms behind this senescence and apoptosis remain unclear. This study aimed to investigate the role of T-box transcription factor T (Tbxt) in IDD both in vitro and in vivo, using a hydrogen peroxide (H2O2)-induced NPCs senescence and apoptosis model, as well as a rat acupuncture IDD model. First, the expression of p16 and cleaved-caspase 3 significantly increased in degenerated human NPCs, accompanied by a decrease in Tbxt expression. Knockdown of Tbxt exacerbated senescence and apoptosis in the H2O2-induced NPCs degeneration model. Conversely, upregulation of Tbxt alleviated these effects induced by H2O2. Mechanistically, bioinformatic analysis revealed that the direct downstream target genes of Tbxt were highly enriched in autophagy-related pathways, and overexpression of Tbxt significantly activated autophagy in NPCs. Moreover, the administration of the autophagy inhibitor, 3-methyladenine, impeded the impact of Tbxt on the processes of senescence and apoptosis in NPCs. Further investigation revealed that Tbxt enhances autophagy by facilitating the transcription of ATG7 through its interaction with a specific motif within the promoter region. In conclusion, this study suggests that Tbxt mitigates H2O2-induced senescence and apoptosis of NPCs by activating ATG7-mediated autophagy.NEW & NOTEWORTHY This study investigates the role of Tbxt in IDD. The results demonstrate that knockdown of Tbxt exacerbates H2O2-induced senescence and apoptosis in NPCs and IDD, whereas upregulation of Tbxt significantly protects against IDD both in vivo and in vitro. Mechanistically, in the nucleus, Tbxt enhances the transcription of ATG7, leading to increased expression of ATG7 protein levels. This, in turn, promotes elevated autophagy levels, ultimately alleviating IDD.

Effect of ADSCs on Th17/Treg and T-bet/GATA-3 in model mice with primary immune thrombocytopenia.

We aimed to observe the effects of adipose-derived mesenchymal stem cells (ADSCs) on T helper 17 (Th17)/regulatory T cells (Treg) and T-box transcription factor (T-bet)/GATA-binding protein 3 (GATA-3) in model mice with primary immune thrombocytopenia (ITP). 32 BALB/C mice were selected. ADSCs were isolated from 2 mice and cultured. The other 30 mice were randomly divided into the normal control group, the ITP model control group, and the ITP experimental group. Platelet count (PLT), Th17/Treg cells, related serum cytokines [interleukin-6 (IL-6), IL-17A, IL-10, and transforming growth factor β1 (TGF-β1)], T-bet and GATA-3 mRNA levels in peripheral blood mononuclear cells (PBMCs) in the 3 groups were detected. PLT and Treg in the ITP experimental group were significantly lower than those in the normal control group (P<0.05), but significantly higher than those in the ITP model control group (P<0.05). Th17 and Th17/Treg in the ITP experimental group were significantly higher than those in the normal control group (P<0.05), but significantly lower than those in the ITP model control group (P<0.05). Serum IL-6 and IL-17A levels, and T-bet mRNA levels in the ITP experimental group were significantly higher than those in the normal control group (P<0.05), but significantly lower than those in the ITP model control group (P<0.05). Serum IL-10 and TGF-β levels, and GATA-3 mRNA levels in the ITP experimental group were significantly lower than those in the normal control group (P<0.05), but significantly higher than those in the ITP model control group (P<0.05). ADSCs can effectively regulate Th17/Treg balance and improve T-bet/GATA-3 mRNA expression levels in ITP model mice.

Programmed cell death-1 is involved with peripheral blood immune cell profiles in patients with hepatitis C virus antiviral therapy.

Mutations in the non-structural protein regions of hepatitis C virus (HCV) are a cause of a non-sustained virological response (SVR) to treatment with direct-acting antivirals (DAAs) for chronic hepatitis; however, there are non-SVR cases without these mutations. In this study, we examined immune cell profiles in peripheral blood before and after ombitasvir/paritaprevir/ritonavir treatment and screened for genes that could be used to predict the therapeutic effects of DAAs. Fluorescence-activated cell sorting analysis indicated that the median frequencies of programmed cell death-1-positive (PD-1+) effector regulatory T cells (eTregs), PD-1+CD8+ T cells, and PD-1+Helper T cells were decreased significantly in SVR cases, but without significant changes in non-SVR cases. The frequency of PD-1+ naïve Tregs was significantly higher in the SVR group than in the non-SVR group before and after treatment. Similar results were found in patients treated with other DAAs (e.g., daclatasvir plus asunaprevir) and supported an immune response after HCV therapy. RNA-sequencing analysis indicated a significant increase in the expression of genes associated with the immune response in the SVR group, while genes related to intracellular and extracellular signal transduction were highly expressed in the non-SVR group. Therefore, we searched for genes associated with PD-1+ eTregs and CD8+ T cells that were significantly different between the SVR and non-SVR groups and found that T-box transcription factor 21 was associated with the non-SVR state. These results indicate that PD-1-related signaling pathways are associated with a non-SVR mechanism after DAAs treatment separate from mutation-related drug resistance.

Cang-ai volatile oil alleviates nasal inflammation via Th1/Th2 cell imbalance regulation in a rat model of ovalbumin-induced allergic rhinitis.

We previously revealed that Cang-ai volatile oil (CAVO) regulates T-cell activity, enhancing the immune response in people with chronic respiratory diseases. However, the effects of CAVO on allergic rhinitis (AR) have not been investigated. Herein, we established an ovalbumin (OVA)-induced AR rat model to determine these effects. Sprague-Dawley (SD) rats were exposed to OVA for 3weeks. CAVO or loratadine (positive control) was given orally once daily for 2weeks to OVA-exposed rats. Behavior modeling nasal allergies was observed. Nasal mucosa, serum, and spleen samples of AR rats were analyzed. CAVO treatment significantly reduced the number of nose rubs and sneezes, and ameliorated several hallmarks of nasal mucosa tissue remodeling: inflammation, eosinophilic infiltration, goblet cell metaplasia, and mast cell hyperplasia. CAVO administration markedly upregulated expressions of interferon-γ, interleukin (IL)-2, and IL-12, and downregulated expressions of serum tumor necrosis factor-α, IL-4, IL-5, IL-6, IL-13, immunoglobulin-E, and histamine. CAVO therapy also increased production of IFN-γ and T-helper type 1 (Th1)-specific T-box transcription factor (T-bet) of the cluster of differentiation-4+ T-cells in splenic lymphocytes, and protein and mRNA expressions of T-bet in nasal mucosa. In contrast, levels of the Th2 cytokine IL-4 and Th2-specific transcription factor GATA binding protein-3 were suppressed by CAVO. These cumulative findings demonstrate that CAVO therapy can alleviate AR by regulating the balance between Th1 and Th2 cells.

Impact of CDK Inhibitors on TBXT Expression in Chordoma Cell Lines Including the First Stable Cell Line of a High-Grade Chordoma.

Chordomas are very rare malignant neoplasms of the bone occurring almost exclusively along the spine. As the tumours are thought to arise from notochordal remnants, the vast majority of chordomas express the TBXT gene, resulting in detectable nuclear amounts of its gene product brachyury. This T-Box transcription factor is commonly recognised as being essential in chordoma cells, and limiting TBXT expression is thought to be the key factor in controlling this tumour. Although the tumour is rare, distinct molecular differences and vulnerabilities have been described with regard to its location and the progression status of the disease, rendering it mandatory for novel cell lines to reflect all relevant chordoma subtypes. Here, we describe a novel chordoma cell line arising from the pleural effusion of a disseminated, poorly differentiated chordoma. This cell line, U-CH22, represents a highly aggressive terminal chordoma and, therefore, fills a relevant gap within the panel of available cell culture models for this orphan disease. CDK7 and CDK9 inhibition was lately identified as being effective in reducing viability in four chordoma cell lines, most likely due to a reduction in brachyury levels. In this study, we determined the capability of the CDK7 inhibitor THZ1 and the CDK1/2/5/9 inhibitor dinaciclib to reduce TBXT expression at mRNA and protein levels in a broad range of nine cell lines that are models of primary, recurrent, and metastasised chordoma of the clivus and the sacrum.

Significant improvement of cardiac outflow tract septation defects in a DiGeorge syndrome model after minoxidil treatment

The T-BOX transcription factor TBX1 is essential for the development of the pharyngeal apparatus and it is haploinsufficient in DiGeorge syndrome (DGS), a developmental anomaly associated with congenital heart disease and other abnormalities. The murine model recapitulates the heart phenotype and showed collagen accumulation.We first used a cellular model to study gene expression during cardiogenic differentiation of WT and Tbx1−/− mouse embryonic stem cells. Then we used a mouse model of DGS to test whether interfering with collagen accumulation using an inhibitor of lysyl hydroxylase would modify the cardiac phenotype of the mutant.We found that loss of Tbx1 in a precardiac differentiation model was associated with up regulation of a subset of ECM-related genes, including several collagen genes. In the in vivo model, early prenatal treatment with Minoxidil, a lysyl hydroxylase inhibitor, ameliorated the cardiac outflow tract septation phenotype in Tbx1 mutant fetuses, but it had no effect on septation in WT fetuses. We conclude that TBX1 suppresses a defined subset of ECM-related genes. This function is critical for OFT septation because the inhibition of collagen cross-linking in the mutant reduces significantly the penetrance of septation defects.