Alantolactone induces concurrent apoptosis and GSDME-dependent pyroptosis of anaplastic thyroid cancer through ROS mitochondria-dependent caspase pathway.

BACKGROUND: Anaplastic thyroid cancer (ATC) is one of the fatal cancers and has not effective treatments. Alantolactone (ATL), a terpenoid extracted from traditional Chinese medicinal herb Inula helenium L., confers significant anti-inflammatory, antibacterial and antitumor activity. However, the activity and mechanisms of ATL in ATC remain unclear. PURPOSE: To investigate the potential anti-ATC effects in vitro and in vivo and the mechanisms involved. METHODS: The anti-proliferative activity of Alantolactone (ATL) against ATC cells was analyzed through CCK-8 and colony formation assays. Flow cytometry assay was performed to assess the cell cycle, cell apoptosis, ROS, and mitochondrial membrane potential (ΔΨm), whereas the cellular localization of cytochrome c and calreticulin were determined using cellular immunofluorescence assays. The lactate dehydrogenase (LDH) enzyme activity in the cell culture medium was measured using a commercial LDH kit, whereas ELISA was conducted to assess the secretory function of IL-1ß. Western blot assays were conducted to determine the expression or regulation of proteins associated with apoptosis and pyroptosis. Subcutaneous tumor model of nude mice was established to evaluate the anticancer activity of ATL in vivo. The expression of Ki67, cyclin B1, cleaved-PARP, cleaved-caspase 3, and IL-1ß in the animal tumor tissues was profiled using immunohistochemistry analyses. RESULTS: Our data showed that ATL significantly inhibited the proliferation and colony formation activity of ATC cells. ATL induced ATC cell cycle arrest at G2/M phase, and downregulated the expression of cyclin B1 and CDC2. Furthermore, ATL induced concurrent apoptosis and pyroptosis in the ATC cells, and the cleavage of PARP and GSDME. It also significantly increased the release of LDH and IL-1ß. Mechanically, ATL-mediated increase in ROS suppressed the Bcl-2/Bax ratio, downregulated the mitochondrial membrane potential and increased the release of cytochrome c, leading to caspase 9 and caspase 3 cleavage. We also found that ATL induced the translocation of an immunogenic cell death marker (calreticulin) to the cell membrane. In addition, it inhibited the growth of the ATC subcutaneous xenograft model, and activated proteins associated with apoptosis and pyroptosis, with a high safety profile. CONCLUSION: Taken together, these results firstly demonstrated that ATL exerted an anti-ATC activity by inducing concurrent apoptosis and GSDME-dependent pyroptosis through ROS-mediated mitochondria-dependent caspase activation. Meanwhile, these cell deaths exhibited obvious characteristics of immunogenic cell death, which may synergistically increase the potential of cancer immunotherapy in ATC. Further studies are needed to explore deeper mechanisms for the anti- ATC activity of ATL.

Lipid-Peptide-mRNA Nanoparticles Augment Radioiodine Uptake in Anaplastic Thyroid Cancer.

Restoring sodium iodide symporter (NIS) expression and function remains a major challenge for radioiodine therapy in anaplastic thyroid cancer (ATC). For more efficient delivery of messenger RNA (mRNA) to manipulate protein expression, a lipid-peptide-mRNA (LPm) nanoparticle (NP) is developed. The LPm NP is prepared by using amphiphilic peptides to assemble a peptide core and which is then coated with cationic lipids. An amphiphilic chimeric peptide, consisting of nine arginine and hydrophobic segments (6 histidine, C18 or cholesterol), is synthesized for adsorption of mRNA encoding NIS in RNase-free conditions. In vitro studies show that LP(R9H6) m NP is most efficient at delivering mRNA and can increase NIS expression in ATC cells by more than 10-fold. After intratumoral injection of NIS mRNA formulated in optimized LPm NP, NIS expression in subcutaneous ATC tumor tissue increases significantly in nude mice, resulting in more iodine 131 (131 I) accumulation in the tumor, thereby significantly inhibiting tumor growth. Overall, this work designs three arginine-rich peptide nanoparticles, contributing to the choice of liposome cores for gene delivery. LPm NP can serve as a promising adjunctive therapy for patients with ATC by restoring iodine affinity and enhancing the therapeutic efficacy of radioactive iodine.

Effect of electroacupuncture and scalp acupuncture combined with language rehabilitation training on cognitive and speech functions of aphasia patients after craniocerebral injury.

OBJECTIVES: To explore the clinical effect of electroacupuncture and scalp acupuncture combined with language rehabilitation training on cognitive and speech functions of patients with aphasia after craniocerebral injury. METHODS: Seventy patients with aphasia after craniocerebral injury from January 2020 and January 2021 were retrospectively collected after filtering. Among them, 35 patients received language rehabilitation training and were included into a control group, and 35 patients who received electroacupuncture and scalp acupuncture combined with language rehabilitation training were included in a study group. Aphasia quotient score, Loewenstein occupational therapy cognitive assessment (LOTCA) score and Boston Diagnostic Aphasia Examination (BDAE) score of the two groups were compared. The relative risk factors of cognitive function and speech function in the patients were explored by multiple regression analysis. RESULTS: The pretreatment aphasia quotient and LOTCA score between the two groups showed no obvious distinction (P>0.05). After treatment, the LOTCA and BDAE scores in the study group were obviously better than those in the control group (P<0.05). CONCLUSIONS: The effect of electroacupuncture and scalp acupuncture combined with language training is remarkable in treating aphasia after severe craniocerebral injury. This combined approach, which can improve the language and cognitive disorders of patients, has valuable application and research prospects in clinic.

Role of oncogene PIM-1 in the development and progression of papillary thyroid carcinoma: Involvement of oxidative stress.

In this study, we aimed to clarify the role of PIM-1 in papillary thyroid carcinoma (PTC) in vitro and investigate the relationship between PIM-1 and redox proteins (NOX4, SOD2, and GPX2) at the tissue and cellular levels. As a PIM-1 inhibitor, SGI-1776 inhibited cell proliferation, colony formation, migration and induced an increase in apoptosis and reactive oxygen species in two PTC cell lines (BCPAP and TPC-1). The expressions of PIM-1, SOD2 and GPX2 were downregulated after siNOX4 exposure. Immunohistochemistry in 120 PTC patients showed that all four proteins exhibited higher expression levels in PTC tissues than in adjacent normal tissues. PIM-1 expression was related to NOX4, SOD2, and GPX2 expressions. The Cancer Genome Atlas database analysis showed the significant correlation between the expression of NOX4 and PIM-1. Our results demonstrated that PIM-1 played an important oncogenic role in PTC carcinogenesis that may be related to oxidative stress.

Predictive Risk-scoring Model For Central Lymph Node Metastasis and Predictors of Recurrence in Papillary Thyroid Carcinoma.

There are about half of papillary thyroid carcinoma (PTC) patients with the experience of central lymph node metastasis (CLNM), while the model to predict high-risk groups of CLNM from PTC patients is uncertain. The aim of this study was to evaluate candidate risk factors of CLNM and identify risk factors of recurrence to guide the postoperative therapeutic decision and follow-up for physicians and patients.A total of 4107 patients(4884 lesions) who underwent lymph node dissection in two hospitals from 2005 to 2014 were evaluated. CLNM risk was stratified and a risk-scoring model was developed on the basis of the identified independent risk factors for CLNM. Cox's proportional hazards regression model was used to investigate the risk factors for recurrence.CLNM was proved in 37.96% (1559/4107) of patients and 33.96% (1659/4884) of lesions. In the multivariate analysis, Male, Age ≤35 years, Tumor size >0.5 cm,Lobe dissemination (+), Psammoma body (+), Multifocality and Capsule invasion (+) were independent risk predictors of CLNM (P < 0.01). A 14-point risk-scoring model was built to predict the stratified CLNM in PTC patients and the area under receiver operating characteristic curve of the model for the prediction of CLNM was 0.672 (95% CI: 0.656-0.688) (P < 0.01). COX regression model showed that Tumor size >0.5 cm, Lobe dissemination (+), Multifocality and CLNM were significant risk factors associated with poor outcomes. The research suggested that prophylactic CLN dissection could be performed in patients with total score ≥4 according to the risk-scoring model, and more aggressive treatment and more frequent follow-up should be considered for patients with Tumor size >0.5 cm, Lobe dissemination (+), Multifocality and CLNM.

Diagnostic values of thyroglobulin in lymph node fine-needle aspiration washout: a systematic review and meta-analysis diagnostic values of FNA-Tg.

Thyroglobulin measurement in the needle washout after fine-needle aspiration (FNA-Tg) served as an important measurement for suspicious recurrent or metastatic lesions. We conducted a pooled analysis to evaluate the diagnostic accuracy of FNA-Tg and searched electronic databases for original articles in English from 1993 through 2017. Finally, a total of 22 studies containing 2,670 lymph nodes (LNs) that enrolled participants with suspicious neck LNs during thyroid nodule workup or papillary thyroid cancer (PTC) follow-up were included. In our analysis, the overall pooled sensitivity for FNA-Tg was 0.91 (95%CI: 0.87-0.93), specificity was 0.94 (95% CI: 0.91-0.96). Meta regression revealed that the cutoff value and status of serum Tg were sources of heterogeneity for sensitivity, and the cutoff value was source of heterogeneity for specificity. Additionally, the cutoff value and status of serum Tg were sources of heterogeneity in the joint model. Subgroup analysis about cut-off value showed that the choice of 1 ng/mL had highest sensitivity, 40 ng/mL had highest specificity. At last, we arrived at the conclusion that FNA-Tg measurement had high specificity and sensitivity in the early detection of LNs metastases from PTC by our meta-analysis. The technique was simple and could be recommended to apply in any FNA facility, especially when LN were small-sized. Significantly, a better standardization of criteria for FNA-Tg detection and cutoff value was required to provide useful data and to improve management of PTC patients in the future.

Clinicopathological and Prognostic Significance of WW Domain Binding Protein 5 Expression in Papillary Thyroid Carcinoma.

OBJECTIVES: Many patients with papillary thyroid cancer (PTC) have a high recurrence risk and poor prognosis, and the main obstacle to the clinical diagnosis and treatment of PTC is lack of effective predictive molecular markers. The purpose of this study was to investigate the clinicopathological and prognostic implications of WW domain binding protein 5 (WBP5) expression in PTC. MATERIALS AND METHODS: Immunohistochemistry of WBP5 was performed using tissue microarrays of 131 patients with PTC who underwent surgery during January 2006 and January 2010 in the Zhejiang Cancer Hospital. Statistical analyses were conducted to evaluate the association between WBP5 expression and the clinicopathological features and to analyze the disease-free survival (DFS) and prognostic factors. RESULTS AND CONCLUSION: The positive expression rate of WBP5 in PTC and the adjacent normal tissues was 42.75% (56/131) and 45.45% (10/22), respectively. WBP5 expression was significantly correlated with bilaterality, capsule invasion, and N-stage, and it was a favorable factor of DFS. Moreover, patients with a high WBP5 expression exhibited reduced risk of disease recurrence compared with that in patients with low WBP5 expression in the univariate analysis, whereas the multivariate analysis suggested that WBP5 was not an independent prognostic factor. Our results indicate that WBP5 might be a favorable prognosis indicator of PTC.

Preliminary screening and analysis of metastasis-related lncRNA and co-expressed papillary thyroid carcinoma mRNA.

The objective of the present study was to investigate the long non-coding RNA (lncRNA) and mRNA expression profiles that are associated with the invasion and metastasis of papillary thyroid carcinoma (PTC). Transwell invasion assays were used to screen three highly invasive sub-strains of the human PTC IHH4 cell line: IHH4-M1, IHH4-M2 and IHH4-M3. In addition, tumor-bearing nude mice were used to identify the invasive and metastatic capacity of the three sub-strains. Agilent lncRNA microarray chips were used to screen 795 differentially expressed lncRNAs and 788 differentially expressed mRNAs. A total of 10 lncRNAs and 10 mRNAs were randomly selected for RT-qPCR validation to confirm that the results were consistent with the microarray chips, suggesting that the results of the microarray chip analysis were relatively accurate. Gene ontology enrichment-based cluster analysis revealed that the differentially expressed genes were mainly associated with steroid biosynthesis, bioadhesion, intercellular adhesion and other metastasis-associated biological processes. The results of the pathway cluster analysis identified that the differentially expressed genes were associated with tumor metastasis-associated signaling pathways, including the cholesterol metabolic signaling pathway, the sterol regulatory element-binding protein signaling pathway and the integrin signaling pathway, suggesting that lncRNA may regulate PTC metastasis through various signaling pathways. The present study screened and constructed PTC metastasis-associated lncRNA and mRNA expression profiles, and it provides a molecular basis for the future study of high-risk molecular markers of PTC.

Expression and prognostic significance of MCM-3 and MCM-7 in salivary adenoid cystic carcinoma.

This study sought to investigate minichromosome maintenance protein 3 (MCM3) and minichromosome maintenance protein 7 (MCM7) expression in salivary adenoid cystic carcinoma (SACC) samples, and to evaluate the relationship between clinicopathological characteristics and prognosis. The expressions of MCM3 and MCM7 were evaluated using immunohistochemistry of tissue sections from SACC patients, and statistical analyses were performed to evaluate the associations between MCM expression and clinicopathological variables and to analyze the disease-free survival (DFS) and prognostic factors. The positive expression rates of MCM3 and MCM7 in SACC were 98.8% and 96.6%, respectively. MCM3 expression correlated with T-stage and nerve invasion. MCM7 expression correlated with T-stage, adjacent tissue invasion, nerve invasion, and prognosis, and was negatively associated with DFS. However, there was no significant correlation between MCM3 expression and DFS. A kappa analysis demonstrated that MCM3 was closely associated with MCM7. MCM7 may be a favorable prognosis indicator in SACC.

Nomograms predicting disease-specific regional recurrence and distant recurrence of papillary thyroid carcinoma following partial or total thyroidectomy.

The study aimed to establish effective nomograms for prediction of tumor regional recurrence and distant recurrence of papillary thyroid carcinoma (PTC) patients after partial or total thyroidectomy.These nomograms were based on a retrospective study on 1034 patients who underwent partial or total thyroidectomy for PTC. The predictive accuracy and discriminative ability of the nomograms were evaluated by the concordance index (C-index) and calibration curve. In addition, a validation cohort was included at the same institution.Multivariate analysis demonstrated that family history, maximal tumor diameter, capsular invasion, and lymph node staging were independent risk factors for regional recurrence-free survival; and family history, histological variants, capsular invasion, perineuronal invasion, and vascular invasion were independent risk factors for distant recurrence-free survival. They were selected into the 2 nomograms, respectively, and the C-index for regional recurrence-free survival and distant recurrence-free survival prediction were 0.72 and 0.83, respectively. In the validation cohort, the 2 nomograms displayed a C-index of 0.72 and 0.89, respectively.The nomograms developed in this study demonstrated their discrimination capability for predicting 3 and 5-year regional recurrence and distant recurrence after partial or total thyroidectomy, and can be used to identify high-risk patients.